Dihydropyridine neuropeptide Y Y1 receptor antagonists 2: Bioisosteric urea replacements

Article abstract of DOI:10.1016/j.bmc.2003.10.016

Structure-activity studies around the urea linkage in BMS-193885 (4a) identified the cyanoguanidine moiety as an effective urea replacement in a series of dihydropyridine NPY Y₁ receptor antagonists. In comparison to urea 4a (K_i=3.3

Full text of DOI:10.1016/j.bmc.2003.10.016

Bioorganic & Medicinal Chemistry 12 (2004) 507–521
Dihydropyridine neuropeptide Y Y1 receptor antagonists 2:
bioisosteric urea replacements
Graham S. Poindexter,* Marc A. Bruce, J. Guy Breitenbucher, Mendi A. Higgins,
S.-Y. Sit, Jeffrey L. Romine, Scott W. Martin, Sally A. Ward, Rachel T. McGovern,
Wendy Clarke, John Russell and Ildiko Antal-Zimanyi
Pharmaceutical Research Institute, Bristol-Myers Squibb Co., 5 Research Parkway, Wallingford, CT, 06492-7660, USA
Received 23 June 2003; accepted 8 October 2003
Abstract—Structure–activity studies around the urea linkage in BMS-193885 (4a) identified the cyanoguanidine moiety as an
effective urea replacement in a series of dihydropyridine NPY Y1 receptor antagonists. In comparison to urea 4a (Ki=3.3 nM),
cyanoguanidine 20 (BMS-205749) displayed similar binding potency at the Y1 receptor (Ki=5.1 nM) and full functional antagon-
ism (Kb=2.6 nM) in SK-N-MC cells. Cyanoguanidine 20 also demonstrated improved permeability properties in Caco-2 cells in
comparison to urea 4a (43 vs 19 nm/s).
# 2003 Elsevier Ltd. All rights reserved.
1. Introduction
have been identified and their inhibitory effects on
feeding described (Fig. 1).7 With respect to the involve-
ment of Y1 receptor antagonists in feeding, BIBP 3226
was the first potent, non-peptidic agent reported with
inhibitory activity on feeding, although the effect
remains controversial.8,9 A more potent and less
controversial analogue, BIBO 3304, was later reported
to attenuate feeding in rats when centrally dosed.10
More recently, several new Y1 antagonists (LY357897,11
J-104870,12 and J-11581413) have appeared which
demonstrate inhibitory effects on feeding in rodents and
support the involvement of the Y1 receptor on food
intake.
Neuropeptide Y (NPY) is a 36-amino acid peptide,
which was first isolated in 1982 from porcine brain.1
The peptide is a member of a larger peptide family
which also includes peptide YY (PYY), pancreatic pep-
tide (PP), and a non-mammalian fish pancreatic peptide
(PY).2 NPY is very highly conserved in a wide variety of
animal, reptile, and fish species and is found in many
central and peripheral sympathetic neurons. NPY is the
most abundant peptide observed in mammalian brain
and has long been implicated in the regulation of feed-
ing behavior and energy homeostasis.3 To date, NPY
remains the most potent orexigenic agent known.
We recently reported structure–activity studies around a
series of novel dihydropyridine NPY Y1 receptor
antagonists which ultimately led to the discovery of
BMS-193885 (4a) (Fig. 1).14 Urea 4a is a potent and
selective Y1 receptor antagonist (Ki=3.3 nM) and dis-
plays anorectic activity in several animal models of
feeding after intraperitoneal (ip) dosing. Although 4a
was found to have satisfactory brain penetrant proper-
ties, its bioavailability after oral administration in rats
was negligible (Fpo=0.1%). Since 4a displayed low per-
meability in Caco-2 cells (Pc=19 nm/s) but showed
good stability after incubation in rat liver microsomes,
it was postulated that its lack of oral exposure was most
likely due to poor intestinal absorption. This was fur-
ther corroborated by ip and intraportal (ipt) dosing
Currently, five different NPY receptor subtypes have
been cloned and characterized (Y1, Y2, Y4, Y5, and y6).4
Although recent evidence suggests the Y5 receptor is
closely associated pharmacologically with the regulation
of food intake, it is generally believed that both Y1 and
Y5 receptors are involved in the feeding response to
NPY.5,6
Over the past several years, a number of specific and
selective, small molecule Y1 and Y5 receptor antagonists
* Corresponding author. Tel.: +1-203-677-6702; fax: +1-203-677-
7702; e-mail: graham.poindexter@bms.com
0968-0896/$ - see front matter # 2003 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2003.10.016

508
G. S. Poindexter et al. / Bioorg. Med. Chem. 12 (2004) 507–521
Figure 1. Known NPY Y1 receptor antagonists.
studies in rats which showed 4a to be completely bio-
available by these routes of administration (Fip=100%,
Fipt=100%).
The physiochemical profile of 4a did not adequately
explain its poor permeability characteristics. Although
the crystalline free base of 4a is poorly soluble in water
(ꢀ2 mg/mL), the l-lactate salt was found to have excel-
lent solubility (3 mg/mLin H O). Except for its mar-
2
ginally high molecular weight (Mr=591) and number of
rotatable bonds (12), the physicochemical properties of
4a15 do not excessively violate the guidelines established
by either Lipinski16 or Veber and Kopple17 for oral
absorption. Structure–activity studies around the che-
motype showed both the dihydropyridine and arylpi-
peridine portions of the molecule were important for
activity, suggesting that reducing Mr and/or the number
of rotatable bonds would be difficult to achieve. Never-
the-less, we felt that improvements could be made on
the general chemotype to increase intestinal perme-
ability and thus enhance oral bioavailability. This
report will describe some of our efforts to find suitable
replacement functionality for the urea linker portion of
BMS-193885 (4a).
2. Results and discussion
We first examined the influence of the urea NH bonds
on both NPY binding affinity and Caco-2 cell perme-
ability. The preparation of urea derivatives 4a, 4b, and 8
are shown in Scheme 1. Dihydropyridine aniline 118 was
treated with excess phosgene in THF to furnish isocyanate
Scheme 1. Synthesis of ureas 4 and 8: (a) COCl2, THF (100%); (b) 3a
or 3b, CH2Cl2 (60% for 4a, 67% for 4b); (c) (CF3CO)2O, pyridine,
CH2Cl2 (74%); (d) NaH, MeI, DMF (90%); (e) NaOH, EtOH (86%);
(f) Cl(CH2)3NCO, CH2Cl2 (75%); (g) 4-(3-methoxyphenyl)piperidine,
NaI, K2CO3, MeCN (46%).

G. S. Poindexter et al. / Bioorg. Med. Chem. 12 (2004) 507–521
509
2. Isocyanate 2 was then condensed in CH2Cl2 with
either aminopropylpiperidine 3a to give parent urea 4a
(BMS-193885), or N-(methylamino)propylpiperidine 3b
to give the corresponding N-methyl derivative 4b.
determined to assess the effect of the selective removal
of the urea and dihydropyridine NH bonds on intestinal
absorption.21 It is thought that amide and amide-like
NH bonds are a major contributor to poor membrane
permeability, due to the desolvation energy required for
a molecule to move from an extracellular aqueous
environment into a hydrophobic cell membrane.22
The isomeric methylated urea 8 was also prepared from
aniline 1 (Scheme 1). Treatment of 1 with trifluoroacetic
anhydride and pyridine in CH2Cl2 gave the corre-
sponding trifluoroacetamide which was subsequently
methylated (NaH/MeI in DMF) to afford N-methyl tri-
fluoroacetamide 5. The amide was hydrolyzed by
refluxing in NaOH/EtOH to yield N-methylaniline 6.
Condensation of 6 with 3-chloropropyl isocyanate in
CH2Cl2 gave urea 7 which, after reaction with 4-(3-
methoxyphenyl)piperidine in MeCN, furnished the
desired N-methyl urea 8.
The Y1 binding and Caco-2 cell results for ureas 4a, 4b,
8, and 12 are summarized in Table 1. In comparison to
the unsubstituted parent 4a, methylation of the distal
NH in 4b enhanced Caco-2 cell permeability (Pc=68
nm/s) but reduced Y1 affinity (Ki=520 nM) suggesting
the removal of one of the hydrogen bond donors and
potential hydration sites had a positive effect on perme-
ability but negative effect on binding. Interestingly,
methylation of the proximal NH in 8 further enhanced
Caco-2 cell permeability (Pc=96 nm/s) but resulted in a
derivative devoid of Y1 activity (Ki=>1000 nM).
Although it appears that removal of the proximal NH
has a profound effect on permeability, it is also clear
that this NH is critical for Y1 receptor recognition.
Similarly, the N-methyl dihydropyridine derivative 12
was less potent than 4a in Y1 binding (Ki=540 nM).
However, methylation of the dihydropyridine ring NH
had little effect on Caco-2 cell permeability (Pc=25 nm/
s) suggesting the urea NH’s play a more influential role in
solvation and intestinal permeability than the former.23
To complete the methylation studies around the
chemotype, N-methyl dihydropyridine 12 was also pre-
pared. As outlined in Scheme 2, N-methyl dihy-
dropyridine 919 was reduced to aniline 10 by
hydrogenolysis using sulfided platinum on carbon, and
the resulting aniline was converted to urethane 11 by
treatment with ClCO2Me in pyridine. The urethane
was subsequently reacted with B-chlorocatecholborane
under the conditions described by Alper20 to give an
intermediate isocyanate. The isocyanate was immedi-
ately condensed with 3a to furnish the desired
N-methyl derivative 12.
All of the compounds were examined in a competition
binding affinity assay using 125I-PYY as the radioligand
in SK-N-MC cell membranes which endogenously
express the human Y1 receptor. Additionally, Caco-2
cell permeability coefficients for these compounds were
Table 1. NPY Y1 binding affinities and Caco-2 cell permeabilities for
ureas 4, 8, and 12
Compd
Urea
NPY Y1
(Ki, nM)a
Caco-2 cell
permeability
(Pc, nm/s)b
4a
3.3 (Æ0.17)c
19
68
96
25
4b (proximal NH)
8 (distal NH)
12
520
>1000
540
a 125I-PYY binding in SK-N-MC cell membranes. Unless otherwise
indicated, the Ki’s were obtained from a single experiment (n=1)
with each point being run in duplicate. Standard reference agent
BIBN 3226 (Fig. 1) displayed a Ki=15 nM in this assay.
b Caco-2 cell permeability’s determined in Biocoat monolayers. The
Pc’s were obtained by averaging the results from two experiments.
For comparison purposes, methoxyinulin and propranolol, the
negative and positive controls, displayed Pc’s of <10 and >100 nm/s
in this assay, respectively.
c The standard error was generated from three curves, each data point
was obtained from the average of six determinations.
Scheme 2. Synthesis of 1-methyldihydropyridine 12: (a) H2, Pt(S)/C,
MeOH, CHCl3 (100%); (b) ClCO2Me, pyridine (74%); (c) B-chloro-
catecholborane, Et3N, THF (100%); (d) 3a, CH2Cl2 (64%).

510
G. S. Poindexter et al. / Bioorg. Med. Chem. 12 (2004) 507–521
Knowing the proximal NH was required for potent Y1
binding affinity, we next prepared a series of urea
replacement derivatives (Table 2) which incorporated
this key structural element. The synthesis of urethane 14
was effected by the condensation isocyanate 2 with
hydroxypropylpiperidine 13 (Scheme 3).
by Tilley26 to give the corresponding cyanoguanidine
20. Interestingly, we observed the monohydrochloride
salt of 20 slowly decomposed on storage at room tem-
perature. The decomposition product was subsequently
identified as the corresponding guanylurea derivative
21. The monohydrochloride salt of 20 was obviously
exposed to small amounts of moisture since hydrolysis
of cyanoguanidines to guanylureas has been observed
by others in low pH environments.26,27 We subsequently
found that salt formation with less acidic acids (e.g.,
maleic acid) afforded stable salt forms. Lastly, the
nitroethylene derivative 24 was prepared from aniline 1
using the method of Young.28 Reaction of 1 with bis
(thiomethyl)nitroethylene gave intermediate 23 which
was subsequently condensed with amine 3a to afford 24.
The preparation of amides 16 and 17 are shown in
Scheme 4. Aniline
1 was treated with diethyl
phosphonoacetic acid, 2-mercaptothiazoline, DCC, and
DMAP using the procedure of Nagoa24 to give the
amido phosphonate 15. After anion formation with
NaH in THF, 15 was condensed with a b-amino alde-
hyde intermediate [prepared from 4-(3-methox-
yphenyl)piperidine and acrolein using the conditions of
Marko25] to furnish a mixture of cis and trans a,b-
unsaturated amides 16a and 16b. The saturated amide
derivative 17 was obtained by reduction of 16a with H2
over Pd/C in MeOH.
The Y1 binding results for urea replacements 14, 16, 17,
20, 21, 22, and 24 are reported in Table 2. Urethane 14
(Ki=480 nM) and the a,b-unsubstituted amide deriva-
tives 16a and 16b (Ki’s=>1000 nM) were considerably
less potent than urea 4a suggesting conformational fac-
tors and possibly the distal urea NH contribute to
binding recognition at the Y1 receptor. Interestingly, the
fully reduced amide 17 showed substantial Y1 binding
potency (Ki=110 nM) in comparison to either urethane
14 or 16a and 16b, but was still 30-fold less potent than
urea 4a, again suggesting the importance of the distal
NH. Whether these results are due to electronic or con-
formational factors is not known at this time.
Cyanoguanidine 21, thiourea 22, and nitroethylene 24
were prepared as shown in Scheme 5. After converting
aniline 1 to the corresponding isothiocyanate 18 by
treatment with thiocarbonyl diimidazole (TCDI) in
DMF, condensation with amine 3a yielded thiourea 22.
Condensation of 18 with sodium cyanamide afforded
the thiolate intermediate 19 which was subsequently
reacted with amine 3a using HgCl2 under the conditions
Scheme 3. Synthesis of urethane 14: (a) CH2Cl2 (61%).
Scheme 4. Synthesis of amides 16 and 17: (a) (EtO)2POCH2CO2H,
DMAP, 2-mercaptothiazoline, DCC, CH2Cl2 (80%); (b) NaH, THF;
(c) acrolein, DBU, 4-(3-methoxyphenyl)piperidine, THF, (15% for
16a, 5% for 16b); (d) H2, 10% Pd/C, MeOH (60%).
Scheme 5. Synthesis of urea replacements 20, 21, 22, and 24: (a)
TCDI, DMF (75%); (b) NaNHCN, EtOH (100%); (c) 3a, HgCl2,
.
THF (58%); (d) 20 HCl, storage at rt; (e) 3a, CHCl3 (100%); (f)
(MeS)2CCHNO2, MeCN (77%); (g) 3a, CH2Cl2 (66%).

G. S. Poindexter et al. / Bioorg. Med. Chem. 12 (2004) 507–521
511
Both the cyanoguanidine 20 (Ki=5.1 nM) and thiourea
22 (Ki=12 nM) derivatives demonstrated potent bind-
ing affinity at the Y1 receptor. These results are not
surprising considering that the bioequivalence of urea,
thiourea, and cyanoguanidine functionalities has been
reported previously in the H2 receptor antagonist
area.27a,28,29 Both guanylurea 21 and nitroethylene 24
were found to be devoid of Y1 activity (Ki’s=>1000
nM). The latter results may possibly be explained by the
increased polar nature of the functionality in compar-
ison to urea 4a, cyanoguanidine 20, and thiourea 22.28
cyclobutene-1,2-dione in DMF to give the intermediate
25, followed by reaction with 3a in DMF. Thiadiazole
oxide 28 was prepared in a similar manner from 1 using
the conditions of Karady.30 Reaction of 1 with 4,5-die-
thoxythiadiazole oxide and AlMe3 in CH2Cl2 furnished
intermediate 27, which was subsequently treated with 3a
to give thiadiazole oxide 28 after chromatography.
Amino-substituted 1,3,4-oxadiazole derivatives 31a and
31b were prepared by condensation of isocyanate 2 in
CH2Cl2 with either hydrazide 29a or 29b to furnish the
respective intermediate acylsemicarbazides 30a and 30b
(Scheme 7). After heating in POCl3, the intermediate
acylsemicarbazides were converted to the respective
aminooxadiazoles 31a and 31b.
We lastly examined a series of heterocycles as urea repla-
cements. As shown in Scheme 6 squaric acid derivative 26
was prepared by treatment of 1 with 3,4-diethoxy-3-
Lastly, imidazolone 34 was prepared according to the
route outlined in Scheme 8. Using the procedure of
Reddy,31 aniline 1 was treated with cyanogen bromide to
give 32 in quantitative yield. Condensation of cyanamide
32 with ethyl glycinate 33 afforded the 4-oxo imidazolone
Table 2. NPY Y1 binding affinities for urea replacements 14, 16, 17,
20, 21, 22, and 24
Compd
Urea replacement
n
NPY Y1 (Ki, nM)a
3.3 (Æ0.17)b
4a
3
14
3
2
2
2
3
480c
>1000c
>1000
110
16a
16b
17
Scheme 6. Synthesis of heterocycles 26 and 28: (a) 3,4-diethoxy-3-
cyclobuten-1,2-dione, DMF (52%); (b) 3a, DMF (54%); (c) Me3Al,
CH2Cl2, 4,5-diethoxythiadiazole oxide (48%); (d) 3a, MeCN (55%).
20
5.1c
21
3
>1000
22
24
3
3
12
>1000
a 125I-PYY binding in SK-N-MC cell membranes. Unless otherwise
indicated, the Ki’s were obtained from a single experiment (n=1)
with each point being run in duplicate. Standard reference agent
BIBN 3226 (Fig. 1) displayed a Ki=15 nM in this assay.
b The standard error was generated from three curves, each data point
was obtained from the average of six determinations.
c The Ki’s were obtained from two experiments (n=2) being run in
duplicate.
Scheme 7. Synthesis of oxadiazoles 31: (a) 2, CH2Cl2 (26% for 30a,
29% for 30b); (b) POCl3 (8% for 31a, 30% for 31b).

512
G. S. Poindexter et al. / Bioorg. Med. Chem. 12 (2004) 507–521
34. The isomeric 5-oxo derivative 35 was prepared in a
similar manner by condensing isothiocyanate 18 with
the sodium salt of propanamine 3a. The resulting thio-
late was then treated with ethyl glycinate and HgCl2 in
THF to give 35.32
the NPY-mediated inhibition of forskolin-stimulated
cAMP accumulation in competitive manner
a
(Kb=2.6Æ0.8 nM), indicating the compound is a full
functional antagonist at the Y1 receptor.34 In additional
competition binding experiments, 20 showed no affinity
(Ki’s>1000 nM) for other cloned human NPY receptor
subtypes (Y2, Y4, and Y5). In comparison to the urea
parent, cyanoguanidine 20 showed improved perme-
ability characteristics in Caco-2 cells (Pc=43 nm/s vs 19
nm/s for 4a). Although this represents only a moderate
enhancement in Caco-2 permeability, the cyanoguanidine
group functions very effectively as a urea replacement in
this series dihydropyridine Y1 receptor antagonists.35
The Y1 binding results for these heterocyclic replace-
ment analogues are reported in Table 3. The two known
urea replacements, squaric acid 26 (Ki=21 nM) and
thiadiazole oxide 28 (Ki=24 nM), demonstrated good
binding affinity at the Y1 receptor, although both were
still less than 10-fold as active compared to parent urea
4a. The oxadiazole derivatives 31a and 31b showed
moderate affinity for the receptor (Ki’s=99 and 160 nM,
respectively), suggesting that the 2-amino-1,3,4-oxadia-
zole moiety can partially function as a urea replacement
in this series of compounds, and that no significant dif-
ference was observed for either the two- or three-carbon
tether. Both of the conformationally restricted, aminoi-
midazolone derivatives 34 and 35 were devoid of Y1
binding affinity (Ki’s=>1000 nM).
In summary, we prepared series of dihydropyridine
derivatives which incorporated both classical and non-
classical urea replacement groups. Out of a series of 12
targeted functionalities, the cyanoguanidine moiety was
identified as the most effective urea replacement. In
comparison to the starting urea 4a, cyanoguanidine
20 (BMS-205749) displayed similar binding potency
at the Y1 receptor and showed improved permeability
Cyanoguanidine 20 was selected for further investiga-
tion based on its potent binding affinity relative to the
parent urea (Ki=5.1 nM vs 3.3 nM for 4a). Scatchard
analysis of 20 based upon the effect of 125I-PYY binding
in SK-N-MC cell membranes, suggested the binding
inhibition is competitive.33 Thus, cyanoguanidine 20 at
a concentration of 10 nM reduced PYY binding affinity
(Kd of 6.5 vs 1.5 nM), without statistically affecting its
binding capacity (Bmax of 4.9Æ0.9 vs 3.5Æ1.1 pmol/mg
protein)]. Functional studies in SK-N-MC cells expres-
sing the human Y1 receptor found that 20 antagonized
Table 3. NPY Y1 binding affinities for heterocycles 26, 28, 31, 34,
and 35
Compd
Heterocycle
n
NPY Y1 (Ki, nM)a
26
3
21b
28
3
24b
31ac
31bc
34
1
2
3
99b
160b
>1000
35
3
>1000b
a 125I-PYY binding in SK-N-MC cell membranes. Unless otherwise
indicated, the Ki’s were obtained from a single experiment (n=1)
with each point being run in duplicate. Standard reference agent
BIBN 3226 (Fig. 1) displayed a Ki=15 nM in this assay.
b The Ki’s were obtained from two experiments (n=2) being run in
duplicate.
Scheme 8. Synthesis of imidazolones 34 and 35: (a) NaHCO3, BrCN,
EtOH (100%); (b) Na2CO3, MeCN (3%); (c) 3a, Na, EtOH; (d) ethyl
glycinate, HgCl2, THF (5%).
c The precursor acylsemicarbazides 30a and 30b displayed Ki’s of 210
and 170 nM, respectively, in the Y1 binding assay.

G. S. Poindexter et al. / Bioorg. Med. Chem. 12 (2004) 507–521
513
properties in Caco-2 cells. Additional examples of
other cyanoguanidine-linked, dihydropyridine Y1
receptor antagonists will be reported in the future.
solid (32.3 g, 96% yield): mp 90–94 ꢁC; IR (KBr) 2246
cmÀ1. This material (31.3 g, 128 mmol) was taken up in
MeOH (850 mL) and 30% aq NH4OH (150 mL) con-
taining Raney Ni and then hydrogenated for 1 h at 50
psi H2 in a Parr apparatus. The catalyst was removed by
filtration through Celite and the filtrate concentrated in
vacuo. The residue was taken up in CH2Cl2, dried over
Na2SO4, and then the volatiles removed in vacuo to
yield a light amber oil. Bulb-to-bulb distillation of the
crude material furnished 26.6 g of 3a as a colorless oil
(84% yield): mp 156–157 ꢁC (maleic acid salt); 1H NMR
(CDCl3) d 7.19 (t, 1H, J=7.5 Hz), 6.79 (d, 1H, J=7.5
Hz), 6.76 (s, 1H), 6.70 (d, 1H, J=7.5 Hz), 3.76 (s, 3H),
3.03 (d, 2H, J=11.4 Hz), 2.73 (t, 2H, J=6.9 Hz), 2.46
(m, 1H), 2.40 (t, 2H, J=7.2 Hz), 1.99 (m, 2H), 1.88 (s,
2H), 1.80 (m, 4H), 1.65 (m, 2H); 13C NMR (CDCl3) d
159.7, 148.2, 129.5, 119.4, 112.7, 111.4, 57.0, 55.3, 54.6,
3. Experimental
3.1. General
Melting points were determined using a Thomas-
Hoover capillary melting apparatus and are uncor-
rected. Elemental analyses were performed at Robert-
son Microlabs, Madison, NJ, USA, and are within
Æ0.4% of calculated values. Unless otherwise indicated,
1H NMR spectra were recorded at 300 MHz and 13C
spectra at 75.5 MHz on a Bruker AC 300 spectrometer
in the indicated solvents. High field NMR spectra were
recorded on a Bruker Avance 500 spectrometer at 500
42.9,
41.0,
33.6,
30.6.
Anal.
calcd
for
1
and 125 MHz for H and 13C, respectively, in the indi-
C15H24N2O 0.5H2O; C, 70.00; H, 9.79; N, 10.89.
Found: C, 70.14; H, 10.01; N, 10.75.
.
cated solvents. IR spectra were recorded on a Perkin-
Elmer Model 1800 FT spectrophotometer. High resolu-
tion mass spectrometry (HRMS) was performed using a
Finnigan MAT 900 spectrometer. Reverse-phase HPLC
purifications were performed using a YMC, Inc.,
20Â250 mm, 5-mm particle size, C18 S5 column: 35–
100% B at a flow rate of 20 mL/min for 20 min and a
gradient time of 20 min. Mobile phase: B=(10%
MeOH, 90% H2O, 0.1% TFA); A=(90% MeOH, 10%
H2O, 0.1% TFA). A Shimadzu SPD-10A detector at
220 nm was used for peak detection. Starting dihy-
dropyridines 118 and 919 and 4-(3-methoxy-
phenyl)piperidine37 were prepared by literature
accounts.
3.1.3. N-Methyl-[4-(3-methoxyphenyl)piperidine-1-propan-
amine (3b). A solution of 3a (1.75 g, 7.06 mmol) and
80 mLof ethyl formate was refluxed overnight (16 h).
After the volatiles were removed in vacuo, and the
crude formamide was taken up in anhydrous THF (40
.
mL) and 24 mmol of Me2S BH3 (12 mLof a 2.0 M
solution in THF) was added. The solution was stirred
16 h at room temperature and then MeOH was carefully
added. After removing the volatiles in vacuo, the resi-
due was taken up in 25 mLof 1 N HCl and refluxed 2 h.
The solution was then made basic with saturated aq
Na2CO3 and extracted with CH2Cl2. The solvent was
removed in vacuo to give 1.4 g of crude 3b as a colorless
oil (ꢀ72% yield). 1H NMR analysis indicated some
N,N-dimethyl propanamine impurity was also present.
3.1.1. 1,4-Dihydro-4-(3-isocyanatophenyl)-2,6-dimethyl-
3,5-pyridinedicarboxylic acid, dimethyl ester (2). A solu-
tion of 1 (27 g, 85 mmol) in anhydrous THF (10 L) was
refluxed under N2 for 30 min to dissolve all solids. After
cooling, the resulting solution was added in dropwise
fashion to 180 mLof a stirred phosgene solution
(1.92 M in toluene) at 0 ꢁC under N2. The resulting
mixture was stirred an additional 30 min while allowing
to warm to room temperature. After sparging with N2
for 16 h to remove any residual phosgene, the volatiles
were removed in vacuo to furnish 31.4 g of the carba-
moyl chloride intermediate as a cream solid. After
standing for 2 days at room temperature, gradual elim-
ination of HCl afforded 28 g of 2 as a cream solid (the
isocyanate was still contaminated with small amounts of
the carbamoyl chloride intermediate): IR (KBr) 2272
3.1.4. 1,4-Dihydro-[3-[[3-[4-(3-methoxyphenyl)-1-piperi-
dinyl]propyl]amino]carbonyl]amino]phenyl]-2,6-dimethyl-
3,5-pyridinedicarboxylic acid, dimethyl ester, L-lactate
salt (4a). A solution of isocyanate 2 (374 mg, 1.1
mmol) in dry CH2Cl2 (20 mL) was added in a dropwise
fashion over 20 min to a stirred solution of the propa-
namine 3a (340 mg, 1.35 mmol) in anhydrous CH2Cl2
(20 mL). The reaction mixture was stirred for 1 h, and
was then rinsed with water (2Â50 mL), and dried
(Na2SO4). After filtration, the solvent was removed in
vacuo to afford a crude residue of 590 mg. The residue
was taken up in MeCN (5 mL) and combined with a
solution of l-lactic acid (90 mg, 1.0 mmol) in MeCN (5
mL), heated, and then allowed to stand overnight. The
resulting white precipitate was collected by filtration,
rinsed with a minimum of MeCN, and recrystallized
from MeCN to furnish 4a as a fluffy white solid (440
mg, 60% yield): mp 125–126 ꢁC; 1H NMR (DMSO-d6) d
8.87 (s, 1H), 8.52 (s, 1H), 7.24 (d,1H, J=8.1 Hz), 7.20
(t, 1H, J=8.0 Hz), 7.10 (s, 1H), 7.02 (t, 1H, J=7.8 Hz),
6.78 (m, 3H), 6.65 (d, 1H, J=7.5 Hz), 6.28 (m, 1H), 4.84
(s, 1H), 3.95 (q, 1H, J=6.9 Hz), 3.73 (s, 3H), 3.55 (s,
6H), 3.10 (m, 4H), 2.50 (m, 3H), 2.25 (s, 6H), 2.18 (m,
2H), 1.67 (m, 6H), 1.21 (d, 3H, J=6.9 Hz); 13C NMR
(DMSO-d6) d 176.9, 167.4, 159.3, 155.3, 148.1, 147.5,
145.6, 140.4, 129.3, 128.2, 119.7, 118.8, 116.4, 115.4,
1
cmÀ1; H NMR (CDCl3) d 7.12 (m, 2H), 6.97 (s, 1H),
6.86 (t, 1H, J=7.5 Hz), 5.80 (s, 1H), 4.99 (s, 1H), 3.66
(s, 6H), 2.36 (s, 6H); 13C NMR (CDCl3) d 168.0, 149.4,
144.7, 133.2, 129.2, 125.5, 124.2, 122.9, 103.6, 51.3, 39.5,
19.8.
3.1.2. 4-(3-Methoxyphenyl)piperidine-1-propanamine (3a).
A solution of 4-(3-methoxyphenyl)piperidine (26.2 g,
137 mmol) and acrylonitrile (10.8 mL, 164 mmol) in
MeCN (150 mL) was refluxed 4 h and then the solvent
was removed in vacuo to yield the b-aminonitrile inter-
mediate as an amber oil. The oil subsequently under-
went a rapid, exothermic crystallization to form a white

514
G. S. Poindexter et al. / Bioorg. Med. Chem. 12 (2004) 507–521
112.4, 111.4, 101.3, 66.0, 55.2, 54.9, 53.3, 50.6, 41.3,
38.4, 37.2, 32.2, 26.6, 20.7, 18.2. Anal. calcd for
C33H42N4O6 C3H6O3 0.5H2O: C, 62.68; H, 7.16; N,
8.12. Found: C, 62.45; H, 7.23; N, 8.21.
allowed to cool to room temperature and extracted with
CH2Cl2. The organic extract was dried (Na2SO4), and
the solvent was removed in vacuo to afford 6 as a pale-
.
.
yellow solid (1.11 g, 86%): mp 182–184 ꢁC; H NMR
1
(DMSO-d6) d 8.78 (s, 1H), 6.91 (t, 1H, J=7.8 Hz), 6.33
(m, 2H), 6.26 (d, 1H, J=8.1 Hz), 5.45 (q, 1H, J=5.1
Hz), 4.82 (s, 1H), 3.56 (s, 6H), 2.60 (d, 3H, J=5.1 Hz),
2.25 (s, 6H); 13C NMR (DMSO-d6) d 169.3, 151.4,
149.9, 147.0, 130.3, 116.2, 112.6, 110.7, 103.3, 52.3, 40.0,
31.4, 19.9. HRMS calcd for C18H22O4NaN2 (M+Na):
353.148. Found: 353.148.
3.1.5. 1,4-Dihydro-[3-[[3-[4-(3-methoxyphenyl)-1-piperi-
dinyl]propyl]methylamino]-carbonyl]amino]phenyl]-2,6-di-
methyl-3,5-pyridinedicarboxylic acid, dimethyl ester,
hydrochloride salt (4b). In a manner similar to that
above for 4a, a solution of 2 (1.40 g, 40.9 mmol) and 3b
(40 mmol) was stirred at room temperature in CH2Cl2
(40 mL) for 2 h. After concentration in vacuo, the resi-
due was purified by flash chromatography (SiO2,
MeOH/CH2Cl2) to furnish 1.05 g (65%) of 4b as a white
solid. The free base was then converted to the hydro-
chloride salt by treatment with ethereal HCl in CH2Cl2
3.1.8. 4-[3-[3-(Chloropropyl)amino]carbonyl]methylami-
no]phenyl]-1,4-dihydro-2,6-dimethyl-3,5-pyridinedicar-
boxylic acid, dimethyl ester (7). A solution of 6 (1.18 g,
3.58 mmol) and 3-chloropropyl isocyanate (0.4 mL, 4
mmol) in CH2Cl2 (100 mL) was refluxed 2 h. After
concentration in vacuo, the solid residue was purified by
chromatography (SiO2, MeOH/CH2Cl2) to afford 1.20 g
and isolated as
a white solid: mp decomposed
(>130 ꢁC); 1H NMR (500 MHz, DMSO-d6) d 10.4 (br s,
1H), 8.91 (s, 1H), 8.24 (s, 1H), 7.25 (m, 3H), 7.05 (t, 1H,
J=8.0 Hz), 6.80 (m, 3H), 6.72 (d, 1H, J=7.5 Hz), 4.87
(s, 1H), 3.70 (s, 3H), 3.57 (m, 2H), 3.55 (s, 6H), 3.40 (t,
2H, J=6.5 Hz), 3.33 (s, 6H), 3.03 (m, 4H), 2.97 (s, 3H),
2.78 (m, 1H), 2.26 (s, 6H), and 2.01 (m, 6H); 13C NMR
(125 MHz, DMSO-d6) d 167.1, 159.3, 155.6, 147.7,
145.7, 145.5, 140.0, 129.5, 127.6, 120.7, 119.0, 118.5,
118.2, 112.5, 111.6, 101.3, 54.9, 53.6, 51.8, 50.5, 40.0,
38.6, 38.4, 34.2, 29.7, 21.9, 18.1. Anal. calcd for
(75%) of 7 as an off-white solid: mp 172–174 ꢁC; H
1
NMR (DMSO-d6) d 7.22 (m, 2H), 7.11 (s, 1H), 7.00 (d,
1H, J=7.3 Hz), 5.87 (s, 1H), 4.99 (s, 1H), 4.53 (t, 2H,
J=5.8 Hz), 3.65 (s, 6H), 3.51 (t, 2H, J=6.4 Hz), 3.28 (q,
2H, J=6.4 Hz), 3.23 (s, 3H), 2.34 (s, 6H), 1.93 (p, 2H,
.
J=6.4 Hz). Anal. calcd for C22H28ClN3O5 0.7H2O: C,
57.13; H, 6.41; N, 9.09. Found: C, 57.08; H, 6.14; N,
9.09.
.
.
C34H44N4O6 HCl H2O: C, 61.95; H, 7.19; N, 8.50.
Found: C, 62.18; H, 7.26; N, 8.44.
3.1.9. 1,4-Dihydro-[3-[[3-[4-(3-methoxyphenyl)-1-piperi-
dinyl]propyl]amino]carbonyl]methylamino]phenyl]-2,6-di-
methyl-3,5-pyridinedicarboxylic acid, dimethyl ester,
hydrochloride salt (8). A mixture of 7 (1.00 g, 2.22
mmol), 4-(3-methoxyphenyl)piperidine (420 mg, 2.2
mmol), K2CO3 (0.35 g, 2.5 mmol), and NaI (25 mg) in
MeCN (50 mL) was refluxed overnight. The mixture
was then partitioned between CH2Cl2 and H2O and the
organic extract dried (Na2SO4), filtered, and con-
centrated in vacuo. The residue was purified by flash
chromatography (SiO2, MeOH/CH2Cl2) to give 610 mg
(46%) of 8. The free base was converted to the hydro-
chloride salt by treatment with ethereal HCl in CH2Cl2
and isolated as a white solid: mp 60–70 ꢁC (sintered); 1H
NMR (500 MHz, DMSO-d6) d 10.5 (br s, 1H), 9.07 (s,
1H), 7.23 (p, 2H, J=7.5 Hz), 7.04 (d, 1H, J=8.0 Hz),
6.98 (m, 2H), 6.80 (m, 3H), 6.23 (br t, 1H), 4.88 (s, 1H),
3.74 (s, 3H), 3.56 (s, 6H), 3.52 (m, 2H), 3.43 (s, 6H), 3.12
(s, 3H), 3.10 (m, 2H), 3.01 (m, 4H), 2.79 (m, 1H), 2.28
(s, 6H), 2.08 (m, 2H), 1.95 (m, 2H), and 1.86 (m, 2H);
13C NMR (125 MHz, DMSO-d6) d 167.3, 159.3, 156.7,
148.7, 145.9, 145.8, 143.5, 129.5, 128.7, 124.2, 124.0,
123.7, 118.5, 112.6, 111.6, 101.1, 54.9, 53.8, 51.7, 50.6,
38.9, 38.7, 38.3, 37.5, 37.0, 29.6, 24.0, 18.1. Anal. calcd
3.1.6.
1,4-Dihydro-2,6-dimethyl-4-[3-[methyl(trifluoro-
acetyl)amino]phenyl]-3,5-pyridinedicarboxylic acid, dime-
thyl ester (5). A solution of 1 (10.0 g, 31.6 mmol) in
CH2Cl2 (500 mL) containing pyridine (2.8 mL, 35
mmol) was cooled to 0 ꢁC. Trifluoroacetic anhydride
(4.7 mL, 33 mmol) was then added dropwise, and the
resulting mixture was stirred at room temperature for 2
h, resulting in a thick suspension. A white solid was
collected by filtration and dried in an Abderhalden
apparatus under reduced pressure for 2 h to afford the
intermediate trifluoroacetamide (9.58 g, 74%). To a
solution of this material (2.06 g, 5.0 mmol) in DMF (20
mL) under N2 was added NaH (60% mineral oil dis-
persion, 240 mg, 6.0 mmol). This mixture was stirred for
10 min, followed by the addition of MeI (0.60 mL, 10
mmol). The reaction mixture was stirred for 1 h, and
then was quenched with H2O (100 mL). A precipitate
readily formed, which was collected by filtration and
rinsed with H2O. The resulting solid was taken up in
CH2Cl2, dried (Na2SO4), and concentrated in vacuo to
furnish 1.91 g (90%) of 5 as a light amber solid: mp
ꢁ
1
184–186 C; H NMR (DMSO-d6) d 8.92 (s, 1H), 7.34
(t, 1H, J=7.8 Hz), 7.19 (m, 2H), 7.09 (s, 1H), 4.87 (s,
1H), 3.52 (s, 6H), 3.25 (s, 3H), 2.26 (s, 6H); 13C NMR
(DMSO-d6) d 197.3, 168.9, 156.9, 151.3, 147.8, 141.4,
131.0, 129.3, 127.8, 126.5, 102.9, 52.3, 41.2, 40.4, 19.8.
.
.
for C34H44N4O6 HCl 3.0H2O: C, 57.74; H, 7.39; N,
8.06. Found: C, 59.11; H, 7.31; N, 7.96.
3.1.10. 4-(3-Aminophenyl)-1,4-dihydro-1,2,6-trimethyl-
3,5-pyridinedicarboxylic acid, dimethyl ester (10). A sus-
pension of 9 (9.26 g, 25.7 mmol), 5% Pt on sulfided
carbon (1.0 g) in a mixture of 2:1 MeOH/CHCl3 (200
mL) was shaken under 65 psi of H2 on a Parr Hydro-
genator overnight (17 h). The mixture was filtered
through Celite and concentrated in vacuo to yield a
solid residue After trituration in a mixture of hot
.
Anal. calcd for C20H21F3N2O5 0.2H2O: C, 55.87; H,
5.02; N, 6.52. Found: C, 55.84; H, 4.96; N, 6.24.
3.1.7. 1,4-Dihydro-2,6-dimethyl-4-[3-(methylamino)phe-
nyl]-3,5-pyridinedicarboxylic acid, dimethyl ester (6). A
solution of 5 (1.67 g, 3.92 mmol) in EtOH (30 mL) and
1 N NaOH (30 mL) was heated to reflux, and then was

G. S. Poindexter et al. / Bioorg. Med. Chem. 12 (2004) 507–521
515
i-PrOH/EtOH, 8.48 g (100%) of 10 was obtained as a
colorless, white solid: mp 213–214 ꢁC; 1H NMR
(DMSO-d6) d 6.79 (t, 1H, J=7.4 Hz), 6.26 (m, 2H), 6.19
(d, 1H, J=7.6 Hz), 4.90 (s, 1H), 4.86 (br s, 1H), 3.60 (s,
6H), 3.15 (s, 3H), and 2.39 (s, 6H); 13C NMR (DMSO-
d6) d 167.7, 149.6, 148.4, 146.1, 128.6, 114.0, 112.2,
111.8, 104.2, 51.0, 37.3, 16.0. Anal. calcd for
115.6, 115.3, 112.7, 111.8, 104.1, 55.0, 54.1, 52.0, 51.1,
39.8, 37.4, 36.5, 34.0, 29.8, 24.6, 16.1. Anal. calcd for
C34H44N4O6 HCl 0.4H2O: C, 62.95; H, 7.12; N, 8.64.
Found: C, 62.95; H, 7.26; N, 8.45.
.
.
3.1.13.
nyl)piperidine (13).
3.1.13.N-3-(Hydroxypropyl)-4-(3-methoxyphe-
mixture of 4-(3-methoxy-
A
.
C18H22N2O4 0.33H2O: C, 64.28; H, 6.79; N, 8.33.
Found: C, 64.29; H, 6.80; N, 8.07.
phenyl)piperidine (2.0 g, 10.4 mmol), 3-bromopropanol
(1.1 mL, 12 mmol), K2CO3 (1.66 g, 12.0 mmol) and
MeCN (50 mL) was stirred at room temperature for 3
days. The solvent was removed in vacuo and the residue
partitioned between CH2Cl2 and H2O. The phases were
separated and the organic portion dried (Na2SO4) and
concentrated. The crude material was purified by flash
chromatography (SiO2, MeOH/CH2Cl2) to give 810 mg
3.1.11. 1,4-Dihydro-4-[3-[(methoxycarbonyl)amino]phe-
nyl]-1,2,6-trimethyl-3,5-pyridinedicarboxylic acid, dime-
thyl ester (11). To a solution of 10 (8.25 g, 25.0 mmol)
in pyridine (50 mL) and CH2Cl2 (50 mL) was slowly
added 2.90 g (27.1 mmol) of methyl chloroformate.
After stirring an additional 3 h at room temperature,
the mixture was poured into water (500 mL) and
extracted with CH2Cl2. The combined organic portions
were washed successively with H2O, 0.1 N HCl, H2O,
and brine and then dried (MgSO4) and concentrated.
The residue was purified by flash chromatography
(SiO2, EtOAc/hexanes) to give 7.20 g (74%) of 11 as a
white solid: mp 163–165 ꢁC; 1H NMR (DMSO-d6) d
9.53 (s, 1H), 7.26 (d, 1H, J=8.1 Hz), 7.17 (s, 1H), 7.08
(t, 1H, J=7.8 Hz), 6.70 (d, 1H, J=7.7 Hz), 4.99 (s, 1H),
3.62 (s, 3H), 3.61 (s, 6H), 3.17 (s, 3H), and 2.41 (s, 6H);
13C NMR (DMSO-d6) d 167.6, 154.0, 150.3, 146.1,
139.2, 128.6, 120.5, 116.4, 115.9, 104.0, 51.6, 51.3, 37.4,
16.0. Anal. calcd for C20H24N2O6: C, 61.85; H, 6.23; N,
7.21. Found: C, 61.88; H, 6.10; N, 7.16.
1
(31%) of 13 as a clear oil: H NMR (CDCl3) d 7.24 (m,
1H), 6.80 (d, 1H, J=7.7 Hz), 6.73 (m, 2H), 3.81 (t, 2H,
J=5.5 Hz), 3.73 (s, 3H), 3.20 (m, 2H), 2.66 (t, 2H,
J=5.7 Hz), 2.50 (m, 1H), 2.07 (m, 2H), 1.79 (m, 6H);
13C NMR (CDCl3) d 159.7, 147.6, 129.4, 119.2, 112.6,
111.5, 64.6, 59.1, 55.2, 54.5, 43.5, 33.3, 27.2. Anal. calcd
.
for C15H23NO2 0.4H2O: C, 70.22; H, 9.35; N, 5.46.
Found: C, 70.18; H, 9.46; N, 5.36.
3.1.14. 1,4-Dihydro-[3-[[3-[4-(3-methoxyphenyl)-1-piperi-
dinyl]propyloxy]carbonyl]amino]phenyl]-2,6-dimethyl-3,5-
pyridinedicarboxylic acid, dimethyl ester, hydrochloride
salt (14). A solution of 2 (656 mg, 1.92 mmol) and 13
(600 mg, 2.41 mmol) in CH2Cl2 (50 mL) was stirred at
room temperature for 2 h and then concentrated in
vacuo. The crude material was purified by flash chro-
matography (SiO2, MeOH/CH2Cl2). The resulting free
base was converted to the hydrochloride salt by treat-
ment with ethereal HCl in CH2Cl2 to give 680 mg (61%)
3.1.12. 1,4-Dihydro-[3-[[3-[4-(3-methoxyphenyl)-1-piperi-
dinyl]propyl]amino]carbonyl]amino]phenyl] - 1,2,6 - trime-
thyl-3,5-pyridinedicarboxylic acid, dimethyl ester,
hydrochloride salt (12). Using the conditions of Alper,20
B-chlorocatecholborane (2.16 g, 14.0 mmol) was added
under N2 in one portion to a solution of 11 (3.90 g, 10.1
mmol) and Et3N (1.60 g, 15.8 mmol) in THF (80 mL).
The resulting mixture was heated with a heat gun for 5
min to effect dissolution and then the volatiles were
removed in vacuo. The residue was taken up in CH2Cl2
and washed successively with 1 N HCl, 1 N NaOH,
brine, and dried (Na2SO4). Filtration and concentration
in vacuo furnished 4.21 g of the intermediate isocyanate
as a gray solid [IR (KBr) 2272 cmÀ1]. The material was
then added to a solution of 3a (2.60 g, 10.5 mmol) in
CH2Cl2 (30 mL) and stirred 3 h at room temperature.
The solution was washed with 1 N HCl, H2O, 0.5 N
NaOH, H2O, brine, and dried (MgSO4). After filtration,
the solution was concentrated and the residue purified
by flash chromatography (SiO2, EtOAc/hexanes) to
afford 3.86 g (64%) of 12 as a yellow solid. The solid
was converted to the hydrochloride salt by treatment
with ethereal HCl and then triturated overnight in ace-
tone/hexane to furnish the salt as a creamy white solid:
of 14 as a white solid: mp 120–132 ꢁC (sintered); H
1
NMR (DMSO-d6) d 10.71 (br s, 1H), 9.63 (s, 1H), 9.00
(s, 1H), 7.23 (m, 3H), 7.13 (m, 1H), 6.83 (m, 4H), 4.88
(s, 1H), 4.17 (t, 2H, J=6.1 Hz), 3.76 (s, 3H), 3.57 (m,
2H), 3.38 (s, 6H), 3.08 (m, 4H), 2.81 (m, 1H), 2.28 (s,
6H), 2.00 (m, 6H); 13C NMR (DMSO-d6) d 167.5, 159.4,
153.3, 148.3, 145.8, 138.6, 129.7, 128.3, 121.2, 118.7,
117.2, 116.0, 112.7, 111.8, 101.2, 61.5, 55.0, 53.3, 52.1,
50.7, 38.4, 29.8, 23.4, 18.2. Anal. calcd for
.
.
C33H41N3O HCl H2O: C, 61.34; H, 6.86; N, 6.50.
Found: C, 61.29; H, 6.85; N, 6.39.
3.1.15. 1,4-Dihydro-4-[3-[[(diethylphosphonato)methyl]-
carbonyl]amino]phenyl]-2,6-dimethyl-3,5-pyridinedicar-
boxylic acid, dimethyl ester (15). Using the procedure of
Nagao,24 a solution of diethyl phosphonoacetic acid
(1.96 g, 10.0 mmol) in CH2Cl2 (30 mL) was treated
sequentially with 4-(dimethylamino)pyridine (DMAP)
(50 mg), 2-mercaptothiazoline (1.19 g, 10.0 mmol), and
1,3-dicyclohexylcarodiimide (DCC) (1.19 g, 10.0 mmol)
at 25 ꢁC. The reaction was stirred for 2 h followed by
the addition of aniline 1 (3.16 g, 10.0 mmol). The mix-
ture was then stirred an additional 90 min, followed by
filtration through Celite. The filtrate was concentrated
in vacuo and the resulting residue was purified by flash
chromatography (SiO2, EtOAc/MeOH) providing 15
(80%) as a gold resin: 1H NMR (CDCl3) d 8.64 (s, 1H),
7.35 (s, 1H), 7.28 (d, 1H, J=7.6 Hz), 7.09 (t, 1H, J=7.6
Hz), 6.98 (d, 1H, J=7.6 Hz), 6.30 (s, 1H), 4.94 (s, 1H),
mp 154–160 ꢁC; H NMR (DMSO-d6) d 10.42 (s, 1H),
1
7.28 (d, 1H, J=8.1 Hz), 7.22 (t, 1H, J=7.5 Hz), 7.07 (s,
1H), 7.01 (t, 1H, J=7.9 Hz), 6.80 (m, 3H), 6.64 (br s,
1H), 6.61 (t, 1H, J=8.0 Hz), 4.96 (s, 1H), 3.71 (s, 3H),
3.60 (s, 6H), 3.51 (m, 2H), 3.18 (s, 3H), 3.05 (m, 6H),
2.76 (m, 1H), 2.41 (s, 6H), 2.06 (m, 2H), and 1.94 (m,
4H); 13C NMR (DMSO-d6) d 167.6, 159.5, 155.7, 150.2,
146.0, 145.9, 140.6, 139.9, 129.7, 128.5, 119.1, 118.7,

516
G. S. Poindexter et al. / Bioorg. Med. Chem. 12 (2004) 507–521
4.10 (m, 4H), 3.58 (s, 6H), 2.90 (d, 2H, J=20.7 Hz),
2.24 (s, 6H), 1.28 (t, 6H, J=7.1 Hz); 13C NMR (CDCl3)
d 167.9, 161.8, 148.2, 144.7, 137.5, 128.3, 123.9, 118.9,
117.8, 103.1, 62.9, 50.8, 39.0, 36.1 (d, J=129 Hz), 19.2,
16.2. Anal. calcd for C23H31N2O8P: C, 55.87; H, 6.32;
N, 5.67. Found: C, 55.51; H, 6.36; N, 5.57.
added 10% Pd on carbon (10 mg). The mixture was
stirred under 1 atm of H2 for 12 h. The mixture was
then filtered through Celite and the filtrate concentrated
in vacuo to provide 122 mg (60%) of 17 as a colorless
white solid: mp 115–120 ꢁC; 1H NMR (CDCl3) d 9.16 (s,
1H), 7.59 (d, 1H, J=8.0 Hz), 7.50 (s, 1H), 7.19 (t, 1H,
J=7.7 Hz), 7.08 (t, 1H, J=8.0 Hz), 6.98 (m, 2H), 6.75
(m, 3H), 4.97 (s, 1H), 3.75 (s, 3H), 3.60 (s, 6H), 3.47 (m,
2H), 2.88 (m, 2H), 2.67 (m, 3H), 2.36 (m, 4H), 2.30 (s,
6H), 1.94 (m, 2H), 1.80 (m, 2H), 1.68 (m, 2H); 13C
NMR (CDCl3) d 170.9, 168.2, 159.7, 148.3, 145.2, 144.5,
138.6, 138.5, 129.7, 128.0, 123.2, 118.8, 117.7, 112.8,
111.8, 102.9, 55.1, 53.0, 52.9, 50.8, 40.2, 38.9, 36.1, 29.9,
23.4, 22.5, 19.2. HRMS calcd for C34H44O6N3 (M+H):
590.3230. Found: 590.3239.
3.1.16. 1,4-Dihydro-4-[3-[[4-[4-(3-methoxyphenyl)-1-pi-
peridinyl]-E-but-1-ene-1-yl]carbonyl]amino]phenyl]-2,6-di-
methyl - 3,5 - pyridinedicarboxylic acid, dimethyl ester,
hydrochloride salt (16a) and 1,4-dihydro-4-[3-[[4-[4-(3-
methoxyphenyl)-1-piperidinyl]-Z-but-1-ene-1-yl]carbony-
l]amino]phenyl] - 2,6 - dimethyl - 3,5 - pyridinedicarboxylic
acid, dimethyl ester, hydrochloride salt (16b). Phospho-
nate 15 (988 mg, 2.00 mmol) in THF (5 mL) was treated
with NaH (240 mg of a 60% suspension in mineral oil,
6.0 mmol) and stirred for 30 min. In a separate flask,
acrolein (133 mL, 2.00 mmol) was added dropwise to a
0 ꢁC solution of 4-(3-methoxyphenyl)piperidine (392
mg, 2.00 mmol), DBU (3.0 mL , 20mmol) and THF (5.0
mL). This mixture was stirred for 20 min. and then added
to the above phosphonate solution. The resulting mix-
ture was stirred for 2 h at 25 ꢁC. The reaction was then
quenched with H2O (10 mL) and the product extracted
with ether/THF (5:1). The residue was concentrated in
vacuo and purified by flash chromatography (SiO2,
EtOAc/MeOH) to afford 172 mg (15%) of 16a and 57 mg
(5%) of 16b as clear oils. Both were converted to their
hydrochloride salts by treatment with ethereal HCl.
3.1.18. 1,4-Dihydro-4-[3-[isothiocyanato]phenyl]-2,6-di-
methyl-3,5-pyridinedicarboxylic acid, dimethyl ester (18).
Aniline 1 (19.0 g, 60.0 mmol) in DMF (250 mL) was
added via addition funnel to a solution of thiocarbo-
nyldiimidazole (TCDI) (11.8 g, 60.0 mmol) in DMF
(100 mL) over 2 h. The reaction was stirred an addi-
tional 30 min and then H2O was added slowly, resulting
in a copious white precipitate. The precipitate was col-
lected by filtration, washed with H2O, and air dried to
afford 16.1 g (75%) of 18 as a white solid: mp 163–
166 ꢁC; IR (KBr) 2122, 2086 cmÀ1; H NMR (DMSO-
1
d6) d 8.98 (s, 1H), 7.31 (t, 1H, J=7.7 Hz), 7.17 (m, 2H),
7.06 (s, 1H), 4.88 (s, 1H), 3.55 (s, 6H), 2.27 (s, 6H); 13C
NMR (DMSO-d6) d 167.1, 149.8, 146.3, 133.2, 129.7,
126.8, 123.9, 123.8, 100.9, 50.8, 38.6, 18.2. Anal. calcd
16a. Mp 155 ꢁC; H NMR (DMSO-d6) d 10.74 (br s,
for C18H18N2O4S 0.5H2O: C, 58.84; H, 5.21; N, 7.63; S,
8.73. Found: C, 58.89; H, 5.04; N, 8.01; S, 8.73.
1
.
1H), 10.12 (s, 1H), 9.01 (s, 1H), 7.51 (d, 1H, J=8.1 Hz),
7.46 (s, 1H), 7.25 (t, 1H, J=8.4 Hz), 7.13 (t, 1H, J=8.1
Hz), 6.82 (m, 4H), 6.72 (d of t, 1H, J=6.6 and 15.4 Hz),
6.26 (d, 1H, J=15.4 Hz), 4.88 (s, 1H), 3.74 (s, 3H), 3.60
(m, 2H), 3.56 (s, 6H), 3.22 (m, 2H), 3.03 (m, 2H), 2.76
(m, 3H), 2.27 (s, 6H), 2.0 (m, 4H); 13C NMR (DMSO-
d6) d 167.4, 162.9, 159.3, 148.1, 145.9, 145.8, 138.8,
138.7, 129.6, 128.3, 126.9, 122.1, 118.6, 118.1, 117.1,
112.7, 111.7, 101.2, 54.9, 54.2, 51.8, 50.6, 38.7, 38.4,
3.1.19. 1,4-Dihydro-4-[3-[(cyanamido)(thiolate)methimi-
no]phenyl]-2,6-dimethyl-3,5-pyridinedicarboxylic acid, di-
methyl ester, sodium salt (19). Cyanamide (2.52 g, 60
mmol) in dry EtOH (50 mL) was added to a solution of
sodium metal (1.38 g, 60 mmol) in EtOH (150 mL).
Isothiocyanate 18 (17.9 g, 50 mmol) in EtOH (150 mL)
was then added to the sodium cyanamide solution via
an addition funnel. After the addition was complete, the
reaction was brought to a brief boil, and then allowed to
stir for 1 h at room temperature. The mixture was then
concentrated and the residue triturated with EtOAc to
.
.
29.7, 26.0, 18.2. Anal. calcd for C34H41N3O6 HCl H2O:
C, 63.60; H, 6.91; N, 6.54. Found: C, 63.49; H, 6.86; N,
6.35.
1
provide 22.6 g (100%) of 19 as a gray solid: H NMR
1
16b. Mp indistinct; H NMR (DMSO-d6) d 10.58 (br
(DMSO-d6) d 9.05 (s, 1H), 7.51 (s, 1H), 7.35 (d, 1H,
J=8.0 Hz), 6.98 (t, 1H, J=8.0 Hz), 6.65 (d, 1H, J=8.0
Hz), 4.84 (s, 1H), 3.57 (s, 6H), 2.27 (s, 6H); 13C NMR
(DMSO-d6) d 186.5, 167.4, 147.4, 145.7, 140.1, 127.4,
121.2, 119.8, 118.7, 117.6, 101.2, 50.6, 38.0, 18.1; IR
(KBr) 2159 cmÀ1. HRMS calcd for C19H19N4O4S:
399.1127. Found: 399.1111.
s, 1H), 10.01 (s, 1H), 8.97 (s, 1H), 7.43 (d, 1H, J=7.8
Hz), 7.36 (s, 1H), 7.25 (t, 1H, J=7.8 Hz), 7.11 (t, 1H,
J=7.8 Hz), 6.80 (m, 4H), 6.07 (m, 1H), 5.80 (m, 1H),
4.87 (s, 1H), 3.74 (s, 3H), 3.70 (m, 2H), 3.55 (s, 6H), 3.20
(m, 2H), 2.97 (m, 2H), 2.77 (m, 2H), 2.26 (m, 6H), 1.99
(m, 4H); 13C NMR (DMSO-d6) d 168.2, 167.4, 159.4,
148.3, 145.8, 145.7, 138.9, 134.8, 129.6, 128.2, 122.0,
121.7, 118.6, 117.9, 116.9, 112.7, 111.7, 101.2, 57.1, 54.9,
51.3, 50.7, 39.9, 39.7, 38.4, 29.7, 18.2. Anal. calcd for
3.1.20. 1,4-Dihydro-4-[3-[[3-[4-(3-methoxyphenyl)-1-piperi-
dinyl]propyl]amino](cyanoimino)methyl]amino]phenyl] - 2,6-
dimethyl-3,5-pyridine dicarboxylic acid, dimethyl ester,
hydrochloride (20). HgCl2 (10.9 g, 40.2 mmol) was
added in 10 portions over 20 min to a solution of 19
(16.9 g, 42.3 mmol) and 3a (14.9 g, 60.1 mmol) in THF
(500 mL). The resulting brown suspension was stirred
an additional 30 min, followed by the addition of H2O
(50 mL). The thick brown material was then filtered
.
.
C34H41N3O6 HCl 2.0H2O: C, 61.86; H, 7.03; N, 6.37;
H2O, 5.52. Found: C, 62.15; H, 6.83; N, 6.00; H2O, 5.60.
3.1.17. 1,4-Dihydro-4-[3-[[4-[4-(3-methoxyphenyl)-1-pi-
peridinyl]butyl]carbonyl]amino]phenyl]-2,6-dimethyl-3,5-
pyridinedicarboxylic acid, dimethyl ester (17). To a solu-
tion of 16a (200 mg, 0.34 mmol) in MeOH (2.0 mL) was

G. S. Poindexter et al. / Bioorg. Med. Chem. 12 (2004) 507–521
517
through Celite, dried (K2CO3), and concentrated in
vacuo. The crude product was purified by flash chro-
matography (SiO2, MeOH/CH2Cl2) to provide 14.4 g
(58%) of 20 as a colorless solid. This material was con-
verted to the hydrochloride salt by treatment with ethe-
(DMSO-d6) d 11.61 (s, 1H), 8.98 (s, 1H), 7.32 (d of d,
1H, J=4.3, 7.9 Hz), 7.12 (m, 3H), 6.77 (s, 1H), 4.92 (s,
1H), 3.57 (s, 6H), 2.46 (s, 3H), 2.29 (s, 6H); 13C NMR
(DMSO-d6) d 167.3, 163.5, 149.0, 146.2, 136.5, 129.1,
126.5, 124.7, 123.9, 107.6, 101.2, 50.9, 38.6, 18.3, 14.5.
Anal. calcd for C20H23N3O6S 0.5H2O: C, 54.29; H,
5.47; N, 9.50; S, 7.25. Found: C, 54.13; H, 5.21; N, 9.29;
S, 7.40.
real HCl: mp 135–140 ꢁC; IR (KBr) 2173 cmÀ1
;
1H
.
NMR (DMSO-d6) d 10.65 (br s, 1H), 9.14 (s, 1H), 9.08
(s, 1H), 7.40 (t, 1H, J=5.3 Hz), 7.23 (m, 2H), 7.03 (m,
2H), 6.92 (d, 1H, J=7.6 Hz), 6.81 (m, 3H), 4.90 (s, 1H),
3.74 (s, 3H), 3.57 (s, 6H), 3.52 (m, 2H), 3.31 (m, 2H), 3.03
(m, 4H), 2.79 (m, 1H), 2.29 (s, 6H), 1.97 (m, 6H); 13C
NMR (DMSO-d6) d 167.3, 159.4, 157.9, 148.5, 146.1,
145.9, 137.4, 129.6, 128.4, 123.3, 121.8, 120.9, 118.6,
117.2, 112.6, 111.7, 101.0, 54.9, 53.6, 51.9, 50.7, 38.2, 29.6,
3.1.24. 1,4-Dihydro-4-[3-[[3-[4-(3-methoxyphenyl)-1-pi-
peridinyl]propyl]amino]-2-(nitro)ethylen-1-yl]amino]phe-
nyl]-2,6-dimethyl-3,5-pyridinedicarboxylic acid, dimethyl
ester, hydrochloride (24). A solution of 23 (868 mg, 2.00
mmol) and 3a (744 mg, 3.00 mmol) in MeOH (20 mL)
was refluxed for 90 min. The resulting solution was
cooled to room temperature and concentrated in vacuo.
The crude material was purified by flash chromato-
graphy (SiO2, MeOH/EtOAc) to afford 835 mg, (66%)
of 24. The product was converted to its hydrochloride
salt by treatment with ethereal HCl: mp 174–184 ꢁC; 1H
NMR (DMSO-d6) d 10.81 (br s, 1H), 9.08 (s, 1H), 7.27
(m, 3H), 7.06 (m, 2H), 6.81 (m, 3H), 5.75 (s, 1H), 4.86
(s, 1H), 3.89 (m, 2H), 3.37 (s, 3H), 3.57 (m, 2H), 3.54 (s,
6H), 3.09 (m, 4H), 2.80 (m, 1H), 2.27 (s, 6H), 2.11 (m,
4H), 1.94 (m, 2H); 13C NMR (DMSO-d6) d 167.4, 159.5,
155.3, 149.6, 148.0, 146.2, 146.0, 129.7, 129.4, 125.7,
122.1, 118.8, 116.2, 112.8, 111.8, 101.3, 98.2, 65.0, 55.1,
53.6, 52.1, 50.9, 38.8, 29.8, 23.4, 18.3, 15.3. Anal. calcd
.
.
23.6, 18.2. Anal. calcd for C34H42N6O5 HCl 2.0H2O: C,
59.44; H, 6.89; N, 12.27. Found: C, 59.33; H, 6.47; N,
12.05.
3.1.21. 4-[3-[(Carbamoylimino)[3-[4-(3-methoxyphenyl)-1-
piperidinyl]propyl]amino]methyl)amino]phenyl]-2,6-dime-
thyl-1,4-dihydropyridine-3,5-dicarboxylic acid dimethyl
ester, trifluoroacetic acid salt (21). The hydrolysis pro-
.
duct was obtained after storage of 20 HCl in a closed
container for several weeks at room temperature. A
small portion of 21 was purified by reverse-phase HPLC
ꢁ
1
and isolated as a white solid: mp 89–93 C; H NMR
(CDCl3) d 7.18–7.08 (m, 4H), 6.91–6.90 (m, 1H), 6.77–
6.69 (m, 4H), 4.97 (s, 1H), 3.75 (s, 3H), 3.58 (s, 6H), 3.34
(m, 2H), 2.94–2.92 (m, 2H), 2.55–2.27 (m, 9H), 1.96 (t,
2H, J=12.0 Hz), 1.72–1.59 (m, 6H), 1.22 (m, 4H); 13C
NMR (CDCl3) d 168.01, 159.57, 147.68, 145.04, 129.36,
129.23, 129.00, 124.33, 122.44, 119.19, 112.81, 111.13,
102.98, 102.69, 55.05, 54.07, 52.99, 52.32, 50.90, 42.47,
39.69, 39.66, 39.19, 32.76, 31.82, 29.66, 29.59, 29.26,
28.26, 26.46, 22.58, 19.20, 19.12, 14.02. HRMS calcd for
C34H45N6O6 (M+H): 633.3300. Found: 633.3420.
.
.
for C34H43N5O7 HCl 2.6H2O: C, 56.96; H, 6.91; N,
9.77. Found: C, 56.66; H, 6.51; N, 9.47.
3.1.25. 4-[3-[(2-Ethoxy-3,4-dioxo-1-cyclobuten-1-yl)ami-
no]phenyl]-1,4-dihydro-2,6-dimethyl-3,5-pyridinedicar-
boxylic acid, dimethyl ester (25). A mixture of 1 (950
mg, 3.01 mmol) and 3,4-diethoxy-3-cyclobutene-1,2-
dione (527 mg, 3.09 mmol) in DMF (6 mL) was heated
to 100 ꢁC for 16 h under N2. After addition of H2O, the
phases were separated and the aqueous portion extrac-
ted with EtOAc (3Â15 mL). After drying the combined
organic extracts (Na2SO4) and concentration in vacuo,
690 mg (52%) of 25 was isolated as off-white, crystalline
solid: mp 249–252 ꢁC; 1H NMR (DMSO-d6) d 10.66 (br
s, 1H), 8.92 (b. s, 1H), 7.21–7.16 (m, 3H), 6.91–6.87 (m,
1H), 4.89 (s, 1H), 4.75 (q, 2H, J=7.1 Hz), 3.55 (s, 6H),
2.27 (s, 6H), 1.41 (t, 3H, J=7.1 Hz); 13C NMR
(DMSO-d6) d 196.9, 184.2, 167.2, 148.7, 145.9, 137.7,
128.4, 122.7, 118.1, 117.1, 100.9, 69.3, 50.5, 18.1, 15.5.
3.1.22. 1,4-Dihydro-[3-[[3-[4-(3-methoxyphenyl)-1-piperi-
dinyl]propyl]amino]thiocarbonyl]amino]phenyl]-2,6-dime-
thyl-3,5-pyridinedicarboxylic acid, dimethyl ester (22). A
mixture of 18 (600 mg, 1.68 mmol) and 3a (420 mg, 1.69
mmol) in CHCl3 (4 mL) was stirred at room tempera-
ture 4 h. The volatiles were removed in vacuo to yield
1
1.01 g (99%) of 22 as a pale-yellow foam: H NMR
(DMSO-d6) d 9.45 (s, 1H), 8.92 (s, 1H), 7.66 (br s, 1H),
7.10 (m, 4H), 6.86 (d, 1H, J=7.5 Hz), 6.80 (m, 3H), 4.87
(s, 1H), 3.72 (s, 3H), 3.54 (s, 6H), 3.47 (m, 2H), 3.95 (br
s, 2H), 2.34 br s, 2H), 2.55 (s, 6H), 1.94 (m, 2H), 1.80
(m, 7H); 13C NMR (DMSO-d6) d 180.0, 167.3, 159.3,
148.3, 147.9, 145.9, 129.2, 128.1, 122.7, 120.5, 118.9,
112.4, 111.3, 101.2, 55.8, 54.9, 53.7, 50.7, 42.6, 41.9,
.
Anal. calcd for C23H24N2O7 0.4H2O: C, 61.71; H, 5.58;
N, 6.26. Found: C, 61.74; H, 5.48; N, 6.37.
3.1.26. 1,4-Dihydro-4-[3-[2-[3-[4-(3-methoxyphenyl)-1-pi-
peridinyl]propyl]amino]-3,4-dioxo-1-cyclobuten-1-yl]ami-
no]phenyl] - 2,3 - dimethyl - 3,5 - pyridindicarboxylic acid,
dimethyl ester (26). A mixture of 25 (54.7 mg, 0.124
mmol) and 3a (41.0 mg, 0.165 mmol) was heated in
DMF (2 mL) at 100 ꢁC for 16 h. After addition of H2O
and cooling to room temperature, the phases were
separated and the aqueous portion extracted with
EtOAc (3Â15 mL). The combined organic portions
were dried (Na2SO4) and then concentrated in vacuo.
The resulting crude material was purified by chromato-
graphy (SiO2, MeOH/CH2Cl2) to give 43 mg (54%) of
38.2,
32.8,
25.7,
18.2.
Anal.
calcd
for
.
C33H42N4SO5 0.47H2O: C, 64.42; H, 7.03; N, 9.11.
Found: C, 64.41; H, 7.16; N, 8.76.
3.1.23. 1,4-Dihydro-4-[3-[1-(thiomethyl)-2-nitroethyl-
ene]amino]phenyl]-2,6-dimethyl-3,5-pyridinedicarboxylic
acid, dimethyl ester (23). Aniline 1 (1.89 g, 5.89 mmol)
and bis(thiomethyl)nitroethylene (0.99 g, 6.00 mmol)
were refluxed in MeCN (15 mL) for 7 h. The reaction
was then concentrated and the residue purified by flash
chromatography (SiO2, MeOH/CH2Cl2) to furnish 2.0 g
(77%) of 23 as a yellow solid: mp 189–191 ꢁC; 1H NMR
1
26 as a white foam: H NMR (DMSO-d6) d 9.86 (br s,

518
G. S. Poindexter et al. / Bioorg. Med. Chem. 12 (2004) 507–521
1H), 8.93 (br s, 1H), 7.95 (br s, 1H), 7.30 (d, 1H, J=7.7
Hz), 7.18–7.13 (m, 4H), 6.83–6.74 (m, 4H), 4.88 (s, 1H),
3.72 (s, 3H), 3.66–3.63 (m, 2H), 3.55 (s, 6H), 3.06 (m,
2H), 2.6–2.4 (m, 2H), 2.26 (s, 6H), 1.9–1.7 (m, 8H); 13C
NMR (DMSO-d6) d 167.4, 159.3, 146.0, 139.0, 129.4,
118.8, 112.5, 101.1, 54.9, 50.7, 18.3.38 Anal. calcd for
dried (MgSO4), filtered and concentrated. The crude
material was subjected to flash chromatography (SiO2,
EtOAc) to afford 7.90 g (74%) the intermediate amino
ester as a yellow oil. A small portion of this material
was converted to the maleic acid salt for characteriza-
tion (mp 103–104 ꢁC). A solution of the free base (7.00
.
.
C36H42N4O7 2.10H2O : C, 63.53; H, 6.84; N, 8.23.
Found: C, 63.48; H, 6.85; N, 8.58.
g, 26.6 mmol), NH2NH2 H2O (2.66 g (83.1 mmol) and
EtOH (75 mL) was refluxed for 24 h. After cooling, the
solution was concentrated and the crude material pur-
ified by flash chromatography (SiO2: ammoniated
MeOH/EtOAc) to afford 6.45 g (92%) of 29a as a col-
3.1.27. (Æ)-1,4-Dihydro-4-[3-[4-(ethoxy)-1,2,5-thiadiaz-
3-ole-1-oxide]amino]phenyl]-2,6-dimethyl-3,5-pyridine-
dicarboxylic acid, dimethyl ester (27). AlMe3 (1.1 mLof
a 2.0 M soln in hexanes; 2.2 mmol) was added to 1
(316 mg, 1.00 mmol) in CH2Cl2 (10 mL) at 0 ꢁC. The
mixture was stirred for 30 min at 0 ꢁC, followed by the
addition of 4,5-diethoxythiadiazole oxide (285 mg, 1.50
mmol). The reaction was then brought to reflux for 18
h, cooled to 0 ꢁC, and quenched by the slow addition of
sat NH4Cl. The crude product was extracted with
CH2Cl2, dried (Na2SO4), and concentrated in vacuo.
The residue was purified by flash chromatography
(SiO2, MeOH/CH2Cl2) to afford 220 mg (48%) of 27 as
orless solid: mp 82–83 ꢁC; H NMR (CDCl3) d 8.22 (br
1
s, 1H), 7.22 (m, 1H), 6.74 (m, 3H), 3.86 (br s, 2H), 3.79
(s, 3H), 3.09 (s, 2H), 2.91 (m, 2H), 2.46 (m, 1H), 2.25
(m, 2H), and 1.45 (m, 4H); 13C NMR (CDCl3) d 170.9,
159.8, 147.6, 129.5, 119.2, 112.8, 111.4, 61.1, 55.2, 55.0,
42.0, and 33.6. Anal. calcd for C14H21N3O2: C, 63.85;
H, 8.04; N, 15.96. Found: C, 63.70; H, 7.96; N, 15.69.
3.1.30.
2-[4-(3-Methoxypheny)-1-piperidinyl]propionic
acid hydrazide (29b). A solution of 4-(3-methoxy-
phenyl)piperidine (5.85 g, 30.6 mmol) and methyl acry-
late (8.9 mL) was stirred at room temperature for 2.5 h.
The volatiles were removed in vacuo and the residue
filtered through a plug of SiO2 to furnish 7.83 g (93%
yield) of the intermediate amino ester as a very pale
yellow oil. A solution of this material (7.20 g, 26.0
ꢁ
1
a yellow solid: mp 191–197 C; H NMR (DMSO-d6) d
10.30 (s, 1H), 8.94 (s, 1H), 7.84 (s, 1H), 7.64 (d, 1H,
J=8.0 Hz), 7.24 (t, 1H, J=8.0 Hz), 6.94 (d, 1H, J=8.0
Hz), 4.93 (s, 1H), 4.55 (m, 2H), 3.56 (s, 6H), 2.27 (s,
6H), 1.45 (t, 3H, J=7.1 Hz); 13C NMR (DMSO-d6) d
167.3, 164.0, 152.6, 148.3, 145.9, 137.7, 128.4, 123.4,
119.5, 118.3, 101.0, 68.9, 50.7, 38.4, 18.2, 13.8. Anal.
.
mmol), NH2NH2 H2O (3.33 g, 104 mmol) and EtOH
(130 mL) was refluxed for 24 h. After cooling, the solu-
tion was concentrated, and the crude material was pur-
ified by flash chromatography (SiO2: ammoniated
MeOH/EtOAc) to give 4.10 g (57%) of 29b as a clear
.
calcd for C21H24N4O6S 0.5H2O: C, 53.72; H, 5.37; N,
11.93. Found C, 53.94; H, 5.25; N, 12.17.
1
3.1.28. (Æ)-1,4-Dihydro-4-[3-[3-[3-[4-(3-methoxyphenyl)-
1-piperidinyl]propyl]amino]-1,2,5-thiadiaz-2-ole-1-oxi-
de]amino]phenyl]-2,6-dimethyl-3,5-pyridinedicarboxylic
acid, dimethyl ester (28). A solution of 27 (263 mg,
0.572 mmol) and 3a (149 mg, 0.601 mmol) in MeCN (3
mL) was stirred at room temperature for 18 h. After
concentration in vacuo, the residue was purified by flash
chromatography (SiO2, MeOH/CH2Cl2) to give 110 mg
(55%) of 28 as a yellow solid: mp 128–134 ꢁC. 1H NMR
(CDCl3) d 7.45 (br s, 1H), 7.11 (t, 1H, J=7.8 Hz), 7.00
(d, 1H, J=7.8 Hz), 6.68 (m, 6H), 6.36 (br s, 1H), 4.97 (s,
1H), 3.70 (s, 3H), 3.54 (s, 6H), 3.29 (m, 1H), 3.19 (m,
1H), 2.98 (m, 2H), 2.42 (m, 3H), 2.26 (s, 3H), 2.19 (s,
3H), 2.08 (m, 2H), 1.75 (m, 6H); 13C NMR (CDCl3) d
168.1, 159.7, 159.2, 153.5, 148.2, 147.4, 145.5, 145.4,
137.7, 129.5, 128.7, 124.4, 119.9, 119.2, 118.1, 112.9,
111.4, 103.1, 102.9, 55.8, 55.2, 54.2, 54.1, 51.2, 51.1,
42.6, 42.2, 38.8, 32.9, 25.1, 19.7, 19.5. Anal. calcd for
oil: H NMR (DMSO-d6) d 9.00 (br s, 1H), 7.19 (t, 1H,
J=7.8 Hz), 6.75 (m, 3H), 4.14 (m, 2H), 3.72 (s, 3H),
2.92 (m, 2H), 2.50 (t, 2H, J=7.1Hz), 2.45 (m, 1H), 2.20
(t, 2H, J=7.1 Hz), 1.97 (m, 2H), and 1.66 (m, 4H).
3.2. General procedure for the preparation of acylsemi-
carbazides 30a and 30b
To a solution of hydrazide 29 (22.8 mmol) in 65 mLof
CH2Cl2, was added isocyanate 2 (23.0 mmol). The
resulting solution was stirred overnight at room tem-
perature, washed with H2O, brine, and dried (MgSO4).
After filtration and concentration in vacuo, the residue
was purified by flash chromatography (SiO2, MeOH/
CH2Cl2).
3.2.1. 1,4-Dihydro-4-[3-[[2-[2-[4-(3-methoxyphenyl)-1-pi-
peridinyl] - 1 - oxoethyl]hydrazino]carbonyl]amino]phenyl -
2,6-dimethyl-3,5-pyridinedicarboxylic acid, dimethyl es-
ter (30a). This compound was obtained as a clear foam
(26% yield): mp indistinct; 1H NMR (DMSO-d6) d 9.51
(br s, 1H), 8.88 (br s, 1H), 8.66 (br s, 1H), 7.88 (br s,
1H), 7.21 (m, 3H), 7.07 (t, 1H, J=7.7 Hz), 6.76 (m, 4H),
4.86 (s, 1H), 3.73 (s, 3H), 3.55 (s, 6H), 3.34 (s, 2H), 3.02
(m, 2H), 2.45 (m, 1H), 2.25 (s, 6H), 2.19 (m, 2H), 1.71
(m, 4H); 13C NMR (DMSO-d6) d 169.4, 167.4, 159.3,
155.2, 148.2, 148.0, 145.6, 139.3, 129.3, 128.3, 120.6,
118.9, 117.1, 116.1, 112.5, 111.3, 101.3, 60.3, 54.9, 53.8,
50.6, 38.4, 32.9, and 18.2. Anal. calcd for
.
C34H42N6O6S 1.9H2O: C, 58.58; H, 6.62; N, 12.05.
Found: C, 58.97; H, 6.32; N, 11.65. HRMS calcd for
C34H43N6O6S (M+H): 663.2965. Found: 663.2976.
3.1.29. 1-[4-(3-Methoxypheny)-1-piperidinyl]acetic acid
hydrazide (29a).
A
mixture of 4-(3-methoxy-
phenyl)piperidine (7.64 g, 40.0 mmol), methyl bro-
moacetate (6.24 g, 40.8 mmol), K2CO3 (6.44 g, 46.7
mmol) and MeCN (125 mL) was refluxed overnight
under N2. After cooling to room temperature, the
volatiles were removed in vacuo and the residue taken
up in H2O and extracted with CH2Cl2. The combined
organic extracts were then washed with H2O, brine,
.
C32H39N5O7 0.81H2O: C, 61.96; H, 6.60; N, 11.29.
Found: C, 61.96; H, 6.38, N, 11.23.

G. S. Poindexter et al. / Bioorg. Med. Chem. 12 (2004) 507–521
519
3.2.2. 1,4-Dihydro-4-[3-[[2-[3-[4-(3-methoxyphenyl)-1-pi-
peridinyl]-1-oxopropyl]hydrazino]carbonyl]amino]phenyl-
2,6-dimethyl-3,5-pyridinedicarboxylic acid, dimethyl es-
ter (30b). This compound was obtained as a creamy
white foam (29% yield): mp indistinct; 1H NMR
(CDCl3) d 8.29 (br s, 1H), 8.16 (br s, 1H), 7.20 (m, 4H),
7.07 (t, 1H, J=7.7 Hz), 6.97 (d, 1H, J=7.9 Hz), 6.74
(m, 4H), 4.97 (s, 1H), 3.78 (s, 3H), 3.59 (s, 6H), 3.01 (m,
2H), 2.60 (m, 2H), 2.43 (m, 3H), 2.20 (s, 6H), 2.03 (m,
2H), and 1.78 (m, 4H); 13C NMR (CDCl3) d 171.6,
168.4, 159.7, 155.3, 148.4, 147.6, 145.4, 138.6, 129.5,
128.5, 122.8, 119.4, 118.8, 117.8, 112.9, 111.4, 103.1,
55.2, 53.6, 51.0, 42.4, 39.0, 33.1, 31.0, and 19.1. Anal.
The crude reaction mixture was purified by column
chromatography (SiO2, EtOAc/hexane) to give 2.90 g
(100%) of 32 as a pale yellow solid: mp 199–201 ꢁC; IR
1
(KBr) 2229 cmÀ1; H NMR (DMSO-d6) d 10.5 (br s.
1H), 9.0 (s, 1H), 7.2 (t, 1H, J=8.0 Hz), 6.8 (m, 2H), 6.7
(d, 1H, J=9.2 Hz), 4.9 (s, 1H), 3.5 (s, 6H), 2.2 (s, 6H);
13C NMR (DMSO-d6) d 167.3, 149.4, 146.0, 138.6,
129.5, 121.2, 113.7, 112.8, 112.3, 101.1, 50.8, 38.5, 18.2.
3.3.4.
3.3.4.N-[4-(3-Methoxyphenyl)-1-piperidinyl]pro-
pyl]glycine, ethyl ester (33). A solution of ethyl bromo-
acetate (1.32 mL, 12.0 mmol) in MeCN (50 mL) was
slowly added over a period of 1 h to a mixture of 3a
(3.00 g, 12.1 mmol) and K2CO3 (8.25 g, 60.0 mmol) in
MeCN (100 mL). After stirring for an additional 1 h at
room temperature, the mixture was filtered through
Celite and the filtrate concentrated in vacuo. The mate-
rial was purified by flash chromatography (SiO2,
MeOH/CH2Cl2) to furnish 1.27 g (32%) of 33 as an oil:
1H NMR (CDCl3) d 7.20 (m, 2H), 6.74 (m, 3H), 4.18 (q,
2H, J=7.2 Hz), 3.78 (s, 3H), 3.40 (s, 2H), 3.11 (m, 2H),
2.68 (t, 2H, J=6.8 Hz), 2.49 (m, 5H), 2.07 (m, 2H), 1.83
.
calcd for C33H41N5O7 0.50H2O: C, 63.05; H, 6.73; N,
11.14. Found: C, 63.04; H, 6.66; N, 11.12.
3.3. General procedure for the preparation of ox-
adiazoles 31a and 31b
A solution of the acylsemicarbazide 30 (3.5 mmol) and
POCl3 (19 mL) was heated on a steam bath until the
reaction was judged complete by TLC analysis (45 min
to 2 h). The dark solution was cooled to room tem-
perature, diluted with CH2Cl2 (100 mL) and then
poured into 300 mLof a stirred ice–water mixture. The
mixture was allowed to warm to room temperature and
then extracted with 1:1 mixture of CH2Cl2 and MeOH.
The combined organic extracts were washed with H2O,
brine, and dried (MgSO4). The volatiles were removed
in vacuo and the residue purified by flash chromato-
graphy (SiO2, EtOAc/MeOH):
1
(M, 5H), 1.26 (t, 3H, J=7.2 Hz); H NMR (CDCl3) d
172.5, 159.5, 147.8, 129.4, 119.3, 112.7, 111.4, 60.8, 57.2,
55.1, 54.3, 50.9, 48.4, 42.6, 33.1, 26.9, 14.2.
3.3.5. 1,4-Dihydro-4-[3-[4,5-dihydro-1-[3-[4-(3-methoxy-
phenyl)-1-piperidinyl]propyl]-4-oxo-1H-imidazol-2-yl]-
aminophenyl]-2,6-dimethyl-3,5-pyridinedicarboxylic acid,
dimethyl ester, trifluoroacetic acid salt (34). A mixture
of 32 (508 mg, 1.49 mmol), 33 (500 mg, 1.50 mmol) and
Na2CO3 (126 mg, 1.50 mmol) in MeCN (25 mL) was
heated at 85 ꢁC for 48 h in a sealed tube. After cooling,
the reaction mixture was concentrated in vacuo and
then prepurified by column chromatography (SiO2,
MeOH/THF/EtOAc). Purification by reverse-phase
preparative HPLC afforded 38.9 mg (3%) of 34 as a
3.3.1. 1,4-Dihydro-4-[3-[5-[4-(3-methoxyphenyl)-1-piperi-
dinyl]methyl]-1,3,4-oxadiazol-2-yl]amino]phenyl]-2,6-di-
methyl - 3,5 - pyridinedicarboxylic acid, dimethyl ester,
hydrochloride salt (31a). The compound was isolated as
an orange-yellow solid (8% yield): mp indistinct; 1H
NMR (DMSO-d6) d 11.2 (br s, 1H), 9.00 (br s, 1H), 7.38
(m, 1H), 7.19 (m, 3H), 6.76 (m, 4H), 4.90 (s, 1H), 3.68
(s, 3H), 3.64 (s, 2H), 3.56 (s, 6H), 2.95 (m, 2H), 2.40
(m,1H), 2.27 (s, 6H), 2.19 (m, 2H), 1.72 (m, 4H). Anal.
1
clear foam: H NMR (500 MHz, MeOD) d 1.86–2.03
(m, 4H), 2.08–2.18 (m, 2H), 2.19 (s, 6H), 3.0–3.15 (m,
3H), 3.15–3.25 (m, 2H), 3.50 (s, 6H), 3.58–3.68 (m, 2H),
3.67 (s, 3H), 3.68–3.70 (m, 2H), 4.91, (s, H), 6.54–7.38
(m, 8H), 8.0 (s, 1H), 8.42 (s, 1). HRMS calcd for
C35H43N5O6 (M+H): 630.3292. Found: 630.3288.
.
.
calcd for C33H37N5O6 2HCl 1.60H2O: C, 56.56; H, 6.06;
N, 9.99. Found: C, 56.56; H, 6.06; N, 9.77.
3.3.2. 1,4-Dihydro-4-[3-[5-[2-[4-(3-methoxyphenyl)-1-pi-
peridinyl]ethyl]-1,3,4-oxadiazol-2-yl]amino]phenyl]-2,6-di-
methyl-3,5-pyridinedicarboxylic acid, dimethyl ester,
hydrochloride salt (31b). The compound was obtained
as a yellow solid (30% yield): mp 75–80 ꢁC (sintered);
13C NMR (DMSO-d6) d 167.3, 160.0, 159.3, 156.3,
148.4, 145.8, 145.7, 138.4, 129.5, 128.6, 120.3, 118.6,
115.9, 114.6, 112.6, 111.6, 101.1, 54.9, 52.6, 51.9, 51.8,
50.6, 38.2, 29.6, 20.0, and 18.1. HRMS calcd for
C33H40N5O6 (M+H): 602.2979. Found: 602.2993.
3.3.6. 1,4-Dihydro-4-[3-[4,5-dihydro-1-[3-[4-(3-methoxy-
phenyl)-1-piperidinyl]propyl]-5-oxo-1H-imidazol-2-yl]-
aminophenyl]-2,6-dimethyl-3,5-pyridinedicarboxylic acid,
dimethyl ester, trifluoroacetic acid salt (35). Sodium
metal (46 mg, 2.0 mmol) was dissolved in abs EtOH (8
mL) and 3a (347 mg, 1.40 mmol) was added. After
being stirred for 10 min, a suspension of 18 (644 mg,
1.88 mmol) in abs EtOH (12 mL) was added, and the
reaction was briefly heated (2 min, heat gun) until the
suspension dissolved. The solution was then stirred an
additional 1 h at room temperature and concentrated in
vacuo. The resulting thiolate was taken up in THF (10
mL) and glycine ethyl ester (247 mg, 2.4 mmol) added.
After cooling to 0 ꢁC, HgCl2 (542 mg, 2 mmol) was
added and the reaction mixture was stirred for 18 h at
room temperature. H2O (1.5 mL) was added and the
solution filtered through Celite and rinsed with EtOH.
The material was purified by reverse-phase preparative
HPLC to isolate 68.7 mg (5%) of 35 as a clear foam:
3.3.3. 4-[3-(Cyanoamino)phenyl]-1,4-dihydro-2,6-dime-
thyl-3,5-pyridinedicarboxylic acid, dimethyl ester (32).
To a suspension of finely powdered dihydropyridine 1
(2.66 g, 8.51 mmol) in EtOH (55 mL) at 0 ꢁC was added
an excess of cyanogen bromide (1.80 g, 17.0 mmol) in a
single portion, followed by the addition of solid
Na2CO3 (2.00 g, 23.8 mmol). The mixture was allowed
to warm to room temperature and stir overnight (16 h).

520
G. S. Poindexter et al. / Bioorg. Med. Chem. 12 (2004) 507–521
1H NMR (500 MHz, MeOD) d 1.95–2.11 (m, 4H), 2.18–
2.27 (m, 2H), 2.32 (s, 6H), 3.09–3.19 (m, 2H), 3.20-3.28
(m, 1H), 3.28–3.34 (m, 2H), 3.62–3.68 (m, 2H), 3.62 (s,
6H), 3.81 (s, 3H), 3.87–3.95 (m, 2H), 4.19 (s, 2H), 5.05
(s, 1H), 6.78–7.51 (m, 8H), 8.0 (s, 1H), 8.55 (s, 1H).
HRMS calcd for C35H43N5O6 (M+H): 630.3292.
Found: 630.3276.
3.5. NPY Y1 functional studies
SK-N-MC cells were seeded on 48-well cell culture
plates (Polyfiltronics catalogue #PF-150-SCC9; ordered
from VWR) at a density of 5Â104–1Â105 cells/mLand
incubated at 37 ꢁC under 5% CO2 overnight. The med-
ium was then aspirated and changed to fresh medium
containing 1 mM IBMX (to inhibit degradation of ade-
nylate cyclase) and 10 mM forskolin (for stimulating
adenylate cyclase) and varying concentrations of NPY
with or without the test compounds. The cell plate was
incubated for 10 min at 37 ꢁC. At the end of the incu-
bation, c-AMP production was terminated by medium
aspiration followed by addition of 1 mL0.1 N HCl. The
plates were subsequently incubated for at least one h at
room temperature before starting radioimmunoassay. c-
AMP levels were measured using a RIA kit from
Amersham (RPA 509) according to the manufacturer’s
instruction. The c-AMP levels were calculated as fmol c-
AMP/well based on a standard curve. Kb’s were calcu-
lated based on Schild-plot analysis. The NPY-induced
inhibition of forskolin-stimulated c-AMP was measured
in absence, and in the presence, of three concentrations
of the test compounds, and dose-ratios were calculated
from IC50 values, and plotted according to Schild.34
3.4. NPY Y1 receptor binding experiments
Membranes were harvested from SK-N-MC cells which
endogenously express the human Y1 receptor. Adherent
cells were washed twice with cold (4 ꢁC) phosphate buf-
fered saline pH 7.4, without Ca2+ and Mg2+. Cells were
then lifted with PBS-based enzyme free cell dissociation
buffer, and pelleted by centrifugation. After the cells
were lysed in ice-cold hypotonic buffer (50 mM HEPES,
5 mM MgCl2 ) using a Dounce homogenizer, they were
pelleted by centrifugation (18,000 RPM, SS-34 rotor, 15
min, 4 ꢁC). The pellets were resuspended, homogenized
in ice-cold hypotonic buffer, and again centrifuged. The
final membrane pellet was resuspended using a 27 G
needle into a small volume of cold (4 ꢁC) hypotonic
buffer (approximately 100 mLper T175 flask, 10–20 mg/
mLprotein concentration). Protein concentration was
measured by the Bradford method using Bio-Rad
Reagent with a BSA standard curve.39 Membranes were
held on ice for up to 2 h or flash frozen in liquid N2 and
stored at À80 ꢁC.
3.6. Caco-2 cell permeability studies
The Caco-2 cell permeability coefficients (Pc’s) for 4a,
4b, 8, 12, and 20 were determined in Biocoat mono-
layers using the procedure previously described by
Chong.41
Membrane solutions were diluted to 0.15 mg/mLin
binding buffer (20 mM HEPES, 137 mM NaCl, 5.4 mM
KH2PO4, 1.26 mM CaCl2 0.81 mM MgSO4) supple-
mented with 1 mg/mLbacitracin, 0.3% BSA, and 10 mg/
mLaprotinin. Test compounds (or PYY for the control
curve) were diluted to 0.1–1.0 mM in DMSO from 10
mM stock solutions. Each of these five concentrations
were then diluted 1:100 into supplemented binding buf-
fer, of which 20 mLwas added to polypropylene assay
tubes containing 30 mLsupplemented binding buffer to
yield a final concentration of 0.1% DMSO. For non-
specific binding, PYY was diluted in 1% DMSO and
Millipore H2O and then added to assay tubes contain-
ing 30 mLsupplemented binding buffer. Duplicate sam-
ples were prepared by mixing 125I PYY (50 mL, 0.1 nM
final concentration), test compounds or competing pep-
tide (PYY) (50 mL, 0.1 nM to 1.0 mM final concen-
tration) or supplemented binding buffer (50 mL0.4%
DMSO for total binding) and finally membrane sus-
pension (100 mL) for a total volume of 200 mL. Samples
were incubated in a 25 ꢁC (Æ2ꢁ) shaking water bath for
1 h. Incubation was terminated by filtration through
Whatman GF/B Filters (pre-soaked in 1% poly-
ethyleneimine for at least 2 h. followed by three 5-mL
washes of ice cold wash buffer (50 mM Tris, pH 7.4).
Samples were then counted for 125I in a Packard Cobra
Auto-Gamma Counter. Non-specific binding was
defined in the presence of 1 mM PYY. Binding data were
analyzed by non-linear regression using ExcelFit or
KaleidaGraph software. The Kd of 125I PYY at the Y1
receptor was 0.35 nM and the maximal binding capacity
0.16 pmol/mg protein. Ki’s were calculated based on the
Cheng–Prusoff equation.40
Acknowledgements
We thank Drs. Stella Huang and John Leet for the
HMBC experiments with 34 and 35 and Dr. Daniel
Schroeder for high field NMR spectra of 4b and 8.
References and notes
1. Tatemoto, K; Carlquist, M.; Mutt, V. Nature 1982, 269,
659.
2. Wahlestedt, C.; Reis, D. J. Ann. Rev. Pharmacol. Toxicol.
1993, 33, 309.
3. (a) Gehlert, D. R. Neuropeptides 1999, 33, 329. (b) Antal-
Zimanyi, I.; Fathi, Z.; Poindexter, G. S. Curr. Pharm.
Des. 1998, 4, 349. (c) Turton, M. D.; O’Shea, D.; Bloom,
S. R. Neuropeptide Y and Drug Develop. 1997, 40, 15.
4. (a) Gehlert, D. R. Proc. Soc. Exp. Biol. Med. 1998, 218, 7.
(b) Blomqvist, A. G.; Herzog, H. Trends Neurosci. 1997,
20, 294.
5. Inui, A. Trends Pharm. Sci. 1999, 20, 43.
6. A recent review has suggested the NPY Y2 receptor might
also be involved in the regulation of energy homeostasis
and satiety: Herzog, H. Drug News Perspt. 2002, 15, 506.
7. (a) Parker, E.; Van Heek, M.; Stamford, A. Eur. J. Phar-
macol. 2002, 440, 173. (b) Kanatani, A.; Ishihara, A.;
Fukami, T.; Ihara, M. Drugs Future 2002, 27, 589. (c)
Antal-Zimanyi, I; Poindexter, G. S. Drug Dev. Res. 2000,
48, 94. (d) Stamford, A. W.; Parker, E. M. Ann. Rep.
Med. Chem. 1999, 34, 31. (e) Gehlert, D. R.; Hipskind,
P. A. Exp. Opin. Invest. Drugs 1997, 6, 1827.

G. S. Poindexter et al. / Bioorg. Med. Chem. 12 (2004) 507–521
521
8. Rudolf, K.; Eberlein, W.; Engel, W.; Wieland, H. A.;
Willim, K. D.; Entzeroth, M.; Wienen, W.; Beck-Sickinger,
A. G.; Doods, H. N. Eur. J. Pharmacol. 1994, 271, 11.
9. Morgan, D. G. A.; Small, C. J.; Abusnana, S.; Turton,
M.; Gunn, I.; Heath, M.; Rossi, M.; Goldstone, A. P.;
O’Shea, D.; Meeran, K.; Smith, D. M.; Bloom, S. Regul.
Pept. 1998, 75-76, 377.
10. (a) Wieland, H. A.; Engel, W.; Eberlein, M.; Rudolf, K.;
Doods, H. N. Brit. J. Pharmacol. 1998, 125, 549. (b)
Doods, H. N.; Engel, W.; Wieland, H. A.; Eberlein, W.;
Rudolf, K.; Judge, M.; Hamilton, B. S. Regul. Pept. 1997,
71, 212. (c) Doods, H. N.; Wieland, H. A.; Engel, W.;
Eberlein, W.; Willim, K.-D.; Entzeroth, M.; Wienen, W.;
Rudolf, K. Regul. Pept. 1996, 65, 71.
11. Hipskind, P. A.; Lobb, K. L.; Nixon, J. A.; Britton, T. C.;
Bruns, R. F.; Catlow, J.; Dieckman-McGinty, D. K.;
Gackenheimer, S. L.; Gitter, B. D.; Iyengar, S.; Schober,
D. A.; Simmons, R. M. A.; Swanson, S.; Zarrinmayeh,
H.; Zimmerman, D. M.; Gehlert, D. R. J. Med. Chem.
1997, 40, 3712.
12. Kanatani, A.; Kanno, T.; Ishihara, A.; Hata, M.; Sakur-
aba, A.; Tanaka, T.; Tsuchiya, Y.; Mase, T.; Fukuroda,
T.; Fukami, T.; Ihara, M. Biochem. Biophys. Res. Com-
mun. 1999, 266, 88.
Chem. 1977, 20, 901. (b) Oyler, A. R.; Naldi, R. E.; Ste-
fanick, S. M.; Lloyd, J. R.; Graden, D. A.; Cotter, M. L.
J. Pharm. Sci. 1989, 78, 21.
28. Young, R. C.; Durant, G. J.; Emmett, J. C.; Ganellin,
C. R.; Graham, M. J.; Mitchell, R. C.; Prain, H. D.;
Roantree, M. L. J. Med. Chem. 1986, 29, 44.
29. Durant, G. J.; Ganellin, C. R.; Hills, D. W.; Miles, P. D.;
Parsons, M. E.; Pepper, E. S.; White, G. R. J. Med.
Chem. 1985, 28, 1414.
30. Karady, S.; Amato, J. S.; Dortmund, D.; Weinstock,
L . M.Heterocycles 1981, 16, 1561.
31. Reddy, N. L.; Hu, L.-Y.; Cotter, R. E.; Fischer, J. B.;
Wong, W. J.; McBurney, R. N.; Weber, E.; Holmes, D. L.;
Wong, S. T.; Prasad, R.; Keana, J. F. W. J. Med. Chem.
1994, 37, 260.
32. Two isomeric products are theoretically possible from
these condensation cyclizations. In both instances it was
reasoned the unsubstituted nitrogen (for 34), and the N-
alkyl substituted nitrogen (for 35), would be more
nucleophilic than the N-aryl nitrogen, and would pre-
ferentially participate in the condensation to furnish the
desired imidazolone products. Thus, as an example in the
formation of 35, isomer i would be preferentially favored
over isomer ii. Heteronuclear multiple bond correlation
(HMBC) analysis confirmed the structural assignments
for both 34 and 35 were correct.
13. Kanatani, A.; Hata, M.; Mashiko, S.; Ishihara, A.; Oka-
moto, O.; Haga, Y.; Ohe, T.; Kanno, T.; Murai, N.; Ishii,
Y.; Fukuroda, T.; Fukami, T.; Ihara, M. Mol. Pharmacol.
2001, 59, 501.
14. Poindexter, G. S.; Bruce, M. A.; LeBoulluec, K. L.;
Monkovic, I.; Martin, S. W.; Parker, E. M.; Iben, L. G.;
McGovern, R. T.; Ortiz, A. A.; Stanley, J. A.; Mattson,
G. K.; Kozlowski, M.; Arcuri, M.; Antal-Zimanyi, I.
Bioorg. Med. Chem. Lett. 2002, 12, 379.
15. Additional physicochemical descriptors for 4a: cLog
P=4.21, cLog D=3.43 (pH 7.4), H-bond donor=3, H-
bond acceptors=5, PSA=120 A . These descriptors were
calculated using Windows XP ACD Software (Version 5).
16. Lipinski, C. A.; Lombardo, F.; Dominy, B. W.; Feeney,
P. J. Adv. Drug Deliv. Rev. 1997, 23, 3.
17. Veber, D. F.; Johnson, S. R.; Cheng, H.-Y.; Smith, B. R.;
Ward, K. W.; Kopple, K. D. J. Med. Chem. 2002, 45, 2615.
18. Bossert, F., Vater, W. US Patent 3,485,847, 1969.
19. Goldmann, S.; Born, L.; Kazda, S.; Pittel, B.; Schram, M.
J. Med. Chem. 1990, 33, 1413.
2
˚
33. Scatchard, G. Ann. N. Y. Acad. Sci. 1949, 51, 660.
34. Schild, H. O. Brit. J. Pharmacol. 1947, 2, 189.
35. Although myriad configurational arrangements of the
cyanoguanidine group are possible, we suggest 20 exists as
a mixture of E,Z and Z,E-isomers based on earlier studies
with similar cyanoguanidine derivatives.27a,36
20. Valli, V. L. K.; Alper, H. J. Org. Chem. 1995, 60, 257.
21. (a) Artursson, P.; Palm, K.; Luthman, K. Drug. Deliv.
Rev. 1996, 22, 67. (b) Kramer, S. D. Pharm. Sci. Tech.
Today 1999, 2, 373.
¨
22. (a) Goodwin, J. T.; Conradi, R. A.; Ho, N. F. H.; Burton,
P. S. J. Med. Chem. 2001, 44, 3721. (b) Conradi, R. A.;
Hilgers, A. R.; Ho, N. F. H.; Burton, P. S. Pharm. Res.
1991, 8, 1453. (c) Karls, M. S.; Rush, B. D.; Wilkinson,
K. F.; Vidmar, T. J.; Burton, P. S.; Ruwart, M. J. Pharm.
Res. 1991, 8, 1477.
23. An alternative explanation to the enhancement of Caco-2
cell permeabilities for the N-methyl urea derivatives 4b
and 8, but not N-methyldihydropyridine 12 could possibly
be due to favorable conformational effects and solute
volume.
36. (a) Ganellin, R. J. Med. Chem. 1981, 24, 913. (b) Pirotte,
B.; Dupont, L.; de Tullio, P.; Masereel, B.; Schynts, M.;
Delarge, J. Helv. Chim. Acta 1993, 76, 1311. (c) Shibata,
M.; Kagawa, K.; Morisaka, K.; Ishida, T.; Inohue, M.
Acta Crystallogr., Sect. C 1983, 39, 1855.
37. Komoto, T.; Hirota, H.; Otsuka, M.; Kotake, J.; Hase-
gawa, S.; Koya, H.; Sato, S.; Sakamoto, T. Chem. Pharm.
Bull. 2000, 48, 1978.
24. Nagao, Y.; Hagiwara, Y.; Yamada, S.; Masahito, O.;
Fujita, E. Chem. Pharm. Bull. 1984, 32, 4686.
25. Chesney, A.; Marko, I. E. Synth. Commun. 1990, 20,
3167.
38. Due to the poor solubility of squarate 26 in DMSO-d6,
not all of the 29 carbon resonances could be differentiated
with confidence from the solvent baseline and hence only
11 are reported.
26. Tilley, J. W.; Ramuz, H. F.; Gerold, M. J. Med. Chem.
1980, 23, 1438.
27. (a) Durant, G. J.; Emmett, C. R.; Ganellin, C. R.; Miles,
P. D.; Parsons, M. E.; Prain, H. D.; White, G. R. J. Med.
39. Bradford, M. Anal. Biochem. 1976, 72, 248.
40. Cheng, M. S.; Prusoff, W. H. Biochemistry 1977, 16, 3310.
41. Chong, S.; Dando, S. A.; Morrison, R. A. Pharm. Res.
1997, 14, 1835.